bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Quorum-Sensing Molecule Farnesol Is a Modulator of Drug Efflux Mediated by ABC Multidrug Transporters and Synergizes with Drugs in Candida albicans
Authors: Sharma Monika , Prasad Rajendra 
Published: July 2011

ABSTRACT

Overexpression of the
CaCDR1
-encoded multidrug efflux pump protein CaCdr1p (
Candida
drug resistance protein 1), belonging to the ATP binding cassette (ABC) superfamily of transporters, is one of the most prominent contributors of multidrug resistance (MDR) in
Candida albicans
. Thus, blocking or modulating the function of the drug efflux pumps represents an attractive approach in combating MDR. In the present study, we provide first evidence that the quorum-sensing molecule farnesol (FAR) is a specific modulator of efflux mediated by ABC multidrug transporters, such as CaCdr1p and CaCdr2p of
C. albicans
and ScPdr5p of
Saccharomyces cerevisiae
. Interestingly, FAR did not modulate the efflux mediated by the multidrug extrusion pump protein CaMdr1p, belonging to the major facilitator superfamily (MFS). Kinetic data revealed that FAR competitively inhibited rhodamine 6G efflux in CaCdr1p-overexpressing cells, with a simultaneous increase in an apparent

K
m

without affecting the
V
max
values and the ATPase activity. We also observed that when used in combination, FAR at a nontoxic concentration synergized with the drugs at their respective nonlethal concentrations, as was evident from their <0.5 fractional inhibitory concentration index (FICI) values and from the drop of 14- to 64-fold in the MIC
80
values in the wild-type strain and in azole-resistant clinical isolates of
C. albicans
. Our biochemical experiments revealed that the synergistic interaction of FAR with the drugs led to reactive oxygen species accumulation, which triggered early apoptosis, and that both could be partly reversed by the addition of an antioxidant. Collectively, FAR modulates drug extrusion mediated exclusively by ABC proteins and is synergistic to fluconazole (FLC), ketoconazole (KTC), miconazole (MCZ), and amphotericin (AMB).
In Vitro
Efficacy of Dicationic Compounds and Mefloquine Enantiomers against Echinococcus multilocularis Metacestodes
Authors: Stadelmann Britta , Küster Tatiana , Scholl Sabrina , Barna Fabienne , Kropf Christian , Keiser Jennifer , Boykin David W. , Stephens Chad E. , Hemphill Andrew 
Published: July 2011

ABSTRACT

The current chemotherapy of alveolar echinococcosis (AE) is based on benzimidazoles such as albendazole and has been shown to be parasitostatic rather than parasiticidal, requiring lifelong duration. Thus, new and more efficient treatment options are urgently needed. By employing a recently validated assay based on the release of functional phosphoglucose isomerase (PGI) from dying parasites, the activities of 26 dicationic compounds and of the (+)- and (−)-
erythro
-enantiomers of mefloquine were investigated. Initial screening of compounds was performed at 40 μM, and those compounds exhibiting considerable antiparasitic activities were also assessed at lower concentrations. Of the dicationic drugs, DB1127 (a diguanidino compound) with activities comparable to nitazoxanide was further studied. The activity of DB1127 was dose dependent and led to severe structural alterations, as visualized by electron microscopy. The (+)- and (−)-
erythro
-enantiomers of mefloquine showed similar dose-dependent effects, although higher concentrations of these compounds than of DB1127 were required for metacestode damage. In conclusion, of the drugs investigated here, the diguanidino compound DB1127 represents the most promising compound for further study in appropriate
in vivo
models for
Echinococcus multilocularis
infection.
Characterization of OXA-181, a Carbapenem-Hydrolyzing Class D β-Lactamase from Klebsiella pneumoniae
Authors: Potron Anaïs , Nordmann Patrice , Lafeuille Emilie , Al Maskari Zaina , Al Rashdi Fatma , Poirel Laurent 
Published: July 2011

ABSTRACT

Klebsiella pneumoniae
KP3 was isolated from a patient transferred from India to the Sultanate of Oman.
K. pneumoniae
KP3 was resistant to all β-lactams, including carbapenems, and expressed the carbapenem-hydrolyzing β-lactamase OXA-181, which differs from OXA-48 by four amino acid substitutions. Compared to OXA-48, OXA-181 possessed a very similar hydrolytic profile. The
bla
OXA-181
gene was located on a 7.6-kb ColE-type plasmid and was linked to the insertion sequence IS
Ecp1
. The IS
Ecp1
-mediated one-ended transposition of
bla
OXA-181
was also demonstrated.
New Transposon Tn
6133
in Methicillin-Resistant Staphylococcus aureus ST398 Contains
vga
(E), a Novel Streptogramin A, Pleuromutilin, and Lincosamide Resistance Gene
Authors: Schwendener Sybille , Perreten Vincent 
Published: July 2011

ABSTRACT

A novel streptogramin A, pleuromutilin, and lincosamide resistance determinant, Vga(E), was identified in porcine methicillin-resistant
Staphylococcus aureus
(MRSA) ST398. The
vga
(E) gene encoded a 524-amino-acid protein belonging to the ABC transporter family. It was found on a multidrug resistance-conferring transposon, Tn
6133
, which was comprised of Tn
554
with a stably integrated 4,787-bp DNA sequence harboring
vga
(E). Detection of Tn
6133
in several porcine MRSA ST398 isolates and its ability to circularize suggest a potential for dissemination.
Cytotoxicity of Voriconazole on Cultured Human Corneal Endothelial Cells
Authors: Han Sang Beom , Shin Young Joo , Hyon Joon Young , Wee Won Ryang 
Published: July 2011

ABSTRACT
The purpose of the present study was to evaluate the toxicity of voriconazole on cultured human corneal endothelial cells (HCECs). HCECs were cultured and exposed to various concentrations of voriconazole (5.0 to 1,000 μg/ml). Cell viability was measured using a Cell Counting Kit-8 (CCK-8) and live/dead viability/cytotoxicity assays. Cell damage was assessed using phase-contrast microscopy after 24 h of exposure to voriconazole. To analyze the effect of voriconazole on the intercellular barrier, immunolocalization of zonula occludens 1 (ZO1) was performed. A flow cytometric assay was performed to evaluate the apoptotic and necrotic effects of voriconazole on HCECs. Cytotoxicity tests demonstrated the dose-dependent toxic effect of voriconazole on HCECs. Voriconazole concentrations of ≥100 μg/ml led to a significant reduction in cell viability. The morphological characteristics of HCECs also changed in a dose-dependent manner. Increasing concentrations of voriconazole resulted in fading staining for ZO1. Higher concentrations of voriconazole resulted in an increased number of propidium iodide (PI)-positive cells, indicating activation of the proapoptotic pathway. In conclusion, voriconazole may have a dose-dependent toxic effect on cultured HCECs. The results of this study suggest that although voriconazole concentrations of up to 50 μg/ml do not decrease cell viability, intracameral voriconazole concentrations of ≥100 μg/ml may increase the risk of corneal endothelial damage.
Molecular Basis for the Selectivity of Antituberculosis Compounds Capreomycin and Viomycin
Authors: Akbergenov Rashid , Shcherbakov Dmitri , Matt Tanja , Duscha Stefan , Meyer Martin , Wilson Daniel N. , Böttger Erik C. 
Published: July 2011

ABSTRACT

Capreomycin and the structurally similar compound viomycin are cyclic peptide antibiotics which are particularly active against
Mycobacterium tuberculosis
, including multidrug resistant strains. Both antibiotics bind across the ribosomal interface involving 23S rRNA helix 69 (H69) and 16S rRNA helix 44 (h44). The binding site of tuberactinomycins in h44 partially overlaps with that of aminoglycosides, and they share with these drugs the side effect of irreversible hearing loss. Here we studied the drug target interaction on ribosomes modified by site-directed mutagenesis. We identified rRNA residues in h44 as the main determinants of phylogenetic selectivity, predict compensatory evolution to impact future resistance development, and propose mechanisms involved in tuberactinomycin ototoxicity, which may enable the development of improved, less-toxic derivatives.
DNA Gyrase Inhibition Assays Are Necessary To Demonstrate Fluoroquinolone Resistance Secondary to
gyrB
Mutations in Mycobacterium tuberculosis
Authors: Pantel Alix , Petrella Stéphanie , Matrat Stéphanie , Brossier Florence , Bastian Sylvaine , Reitter Delphine , Jarlier Vincent , Mayer Claudine , Aubry Alexandra 
Published: July 2011

ABSTRACT

The main mechanism of fluoroquinolone (FQ) resistance in
Mycobacterium tuberculosis
is mutation in DNA gyrase (GyrA
2
GyrB
2
), especially in
gyrA
. However, the discovery of unknown mutations in
gyrB
whose implication in FQ resistance is unclear has become more frequent. We investigated the impact on FQ susceptibility of eight
gyrB
mutations in
M. tuberculosis
clinical strains, three of which were previously identified in an FQ-resistant strain. We measured FQ MICs and also DNA gyrase inhibition by FQs in order to clarify the role of these mutations in FQ resistance. Wild-type GyrA, wild-type GyrB, and mutant GyrB subunits produced from engineered
gyrB
alleles by mutagenesis were overexpressed in
Escherichia coli
, purified to homogeneity, and used to reconstitute highly active gyrase complexes. MICs and DNA gyrase inhibition were determined for moxifloxacin, gatifloxacin, ofloxacin, levofloxacin, and enoxacin. We demonstrated that the eight substitutions in GyrB (D473N, P478A, R485H, S486F, A506G, A547V, G551R, and G559A), recently identified in FQ-resistant clinical strains or encountered in
M. tuberculosis
strains isolated in France, are not implicated in FQ resistance. These results underline that, as opposed to phenotypic FQ susceptibility testing, the DNA gyrase inhibition assay is the only way to prove the role of a DNA gyrase mutation in FQ resistance. Therefore, the use of FQ in the treatment of tuberculosis (TB) patients should not be ruled out only on the basis of the presence of mutations in
gyrB
.
Salmonella enterica Serovar Typhimurium Colonizing the Lumen of the Chicken Intestine Grows Slowly and Upregulates a Unique Set of Virulence and Metabolism Genes
Authors: Harvey P. C. , Watson M. , Hulme S. , Jones M. A. , Lovell M. , Berchieri A. , Young J. , Bumstead N. , Barrow P. 
Published: July 2011

ABSTRACT

The pattern of global gene expression in
Salmonella enterica
serovar Typhimurium bacteria harvested from the chicken intestinal lumen (cecum) was compared with that of a late-log-phase LB broth culture using a whole-genome microarray. Levels of transcription, translation, and cell division
in vivo
were lower than those
in vitro
.
S.
Typhimurium appeared to be using carbon sources, such as propionate, 1,2-propanediol, and ethanolamine, in addition to melibiose and ascorbate, the latter possibly transformed to
d
-xylulose. Amino acid starvation appeared to be a factor during colonization. Bacteria in the lumen were non- or weakly motile and nonchemotactic but showed upregulation of a number of fimbrial and
Salmonella
pathogenicity island 3 (SPI-3) and 5 genes, suggesting a close physical association with the host during colonization.
S.
Typhimurium bacteria harvested from the cecal mucosa showed an expression profile similar to that of bacteria from the intestinal lumen, except that levels of transcription, translation, and cell division were higher and glucose may also have been used as a carbon source.
Pneumocystis carinii Interactions with Lung Epithelial Cells and Matrix Proteins Induce Expression and Activity of the PcSte20 Kinase with Subsequent Phosphorylation of the Downstream Cell Wall Biosynthesis Kinase PcCbk1
Authors: Kottom Theodore J. , Burgess Joshua W. , Limper Andrew H. 
Published: July 2011

ABSTRACT

Eukaryotic cell proliferation and phenotype are highly regulated by contact-dependent mechanisms. We have previously shown that the binding and interaction of the opportunistic fungal pathogen
Pneumocystis carinii
to lung epithelial cells and extracellular matrix proteins induces mRNA expression of both the mitogen-activated protein (MAP) kinase
P. carinii
Ste20 (PcSte20) and the cell wall-remodeling enzyme
PcCbk1
(16). Herein, we report that in addition to
PcSte20
mRNA expression being upregulated,
Pneumocystis
PcSte20 kinase activity is increased upon interacting with these same lung targets. This activity is also significantly suppressed by
Clostridium difficile
toxin B, a pan-specific inhibitor of small GTPases, demonstrating the potential role of a Cdc42-like molecule in this signaling cascade. We further observed that the PcSte20 kinase physically interacts with a specific region of the
P. carinii
cell wall biosynthesis kinase, PcCbk1, a downstream kinase important for mating projection formation and cell wall remodeling. This direct binding was mapped to a specific region of the PcCbk1 protein. We also demonstrated that PcSte20 obtained from whole
P. carinii
lysates has the ability to phosphorylate PcCbk1 after the organism interacts with lung epithelial cells and extracellular matrix components. These observations provide new insights into
P. carinii
signaling induced by interactions of this important opportunistic fungal pathogen with lung epithelial cells and matrix.
Protective Live Oral Brucellosis Vaccines Stimulate Th1 and Th17 Cell Responses
Authors: Clapp Beata , Skyberg Jerod A. , Yang Xinghong , Thornburg Theresa , Walters Nancy , Pascual David W. 
Published: July 2011

ABSTRACT

Zoonotic transmission of brucellosis often results from exposure to
Brucella
-infected livestock, feral animals, or wildlife or frequently via consumption of unpasteurized milk products or raw meat. Since natural infection of humans often occurs by the oral route, mucosal vaccination may offer a means to confer protection for both mucosal and systemic tissues. Significant efforts have focused on developing a live brucellosis vaccine, and deletion of the
znuA
gene involved in zinc transport has been found to attenuate
Brucella abortus
. A similar mutation has been adapted for
Brucella melitensis
and tested to determine whether oral administration of Δ
znuA
B. melitensis
can confer protection against nasal
B. melitensis
challenge. A single oral vaccination with Δ
znuA
B. melitensis
rapidly cleared from mice within 2 weeks and effectively protected mice upon nasal challenge with wild-type
B. melitensis
16M. In 83% of the vaccinated mice, no detectable brucellae were found in their spleens, unlike with phosphate-buffered saline (PBS)-dosed mice, and vaccination also enhanced the clearance of brucellae from the lungs. Moreover, vaccinated gamma interferon-deficient (IFN-γ
−/−
) mice also showed protection in both spleens and lungs, albeit protection that was not as effective as in immunocompetent mice. Although IFN-γ, interleukin 17 (IL-17), and IL-22 were stimulated by these live vaccines, only RB51-mediated protection was codependent upon IL-17 in BALB/c mice. These data suggest that oral immunization with the live, attenuated Δ
znuA
B. melitensis
vaccine provides an attractive strategy to protect against inhalational infection with virulent
B. melitensis
.
Type 1 Fimbrial Adhesin FimH Elicits an Immune Response That Enhances Cell Adhesion of Escherichia coli
Authors: Tchesnokova Veronika , Aprikian Pavel , Kisiela Dagmara , Gowey Sarah , Korotkova Natalia , Thomas Wendy , Sokurenko Evgeni 
Published: July 2011

ABSTRACT

Escherichia coli
causes about 90% of urinary tract infections (UTI), and more than 95% of all UTI-causing
E. coli
express type 1 fimbriae. The fimbrial tip-positioned adhesive protein FimH utilizes a shear force-enhanced, so-called catch-bond mechanism of interaction with its receptor, mannose, where the lectin domain of FimH shifts from a low- to a high-affinity conformation upon separation from the anchoring pilin domain. Here, we show that immunization with the lectin domain induces antibodies that exclusively or predominantly recognize only the high-affinity conformation. In the lectin domain, we identified four high-affinity-specific epitopes, all positioned away from the mannose-binding pocket, which are recognized by 20 separate clones of monoclonal antibody. None of the monoclonal or polyclonal antibodies against the lectin domain inhibited the adhesive function. On the contrary, the antibodies enhanced FimH-mediated binding to mannosylated ligands and increased by severalfold bacterial adhesion to urothelial cells. Furthermore, by natural conversion from the high- to the low-affinity state, FimH adhesin was able to shed the antibodies bound to it. When whole fimbriae were used, the antifimbrial immune serum that contained a significant amount of antibodies against the lectin domain of FimH was also able to enhance FimH-mediated binding. Thus, bacterial adhesins (or other surface antigens) with the ability to switch between alternative conformations have the potential to induce a conformation-specific immune response that has a function-enhancing rather than -inhibiting impact on the protein. These observations have implications for the development of adhesin-specific vaccines and may serve as a paradigm for antibody-mediated enhancement of pathogen binding.
Membrane Translocation of Binary Actin-ADP-Ribosylating Toxins from Clostridium difficile and Clostridium perfringens Is Facilitated by Cyclophilin A and Hsp90
Authors: Kaiser Eva , Kroll Claudia , Ernst Katharina , Schwan Carsten , Popoff Michel , Fischer Gunter , Buchner Johannes , Aktories Klaus , Barth Holger 
Published: July 2011

ABSTRACT

Some hypervirulent strains of
Clostridium difficile
produce the binary actin-ADP-ribosylating toxin
C. difficile
transferase (CDT) in addition to Rho-glucosylating toxins A and B. It has been suggested that the presence of CDT increases the severity of
C. difficile
-associated diseases, including pseudomembranous colitis. CDT contains a binding and translocation component, CDTb, that mediates the transport of the separate enzyme component CDTa into the cytosol of target cells, where CDTa modifies actin. Here we investigated the mechanism of cellular CDT uptake and found that bafilomycin A1 protects cultured epithelial cells from intoxication with CDT, implying that CDTa is translocated from acidified endosomal vesicles into the cytosol. Consistently, CDTa is translocated across the cytoplasmic membranes into the cytosol when cell-bound CDT is exposed to acidic medium. Radicicol and cyclosporine A, inhibitors of the heat shock protein Hsp90 and cyclophilins, respectively, protected cells from intoxication with CDT but not from intoxication with toxins A and B. Moreover, both inhibitors blocked the pH-dependent membrane translocation of CDTa, strongly suggesting that Hsp90 and cyclophilin are crucial for this process. In contrast, the inhibitors did not interfere with the ADP-ribosyltransferase activity, receptor binding, or endocytosis of the toxin. We obtained comparable results with the closely related iota-toxin from
Clostridium perfringens
. Moreover, CDTa and Ia, the enzyme component of iota-toxin, specifically bound to immobilized Hsp90 and cyclophilin A
in vitro
. In combination with our recently obtained data on the C2 toxin from
C. botulinum
, these results imply a common Hsp90/cyclophilin A-dependent translocation mechanism for the family of binary actin-ADP-ribosylating toxins.
Effect of Deletion of Genes Involved in Lipopolysaccharide Core and O-Antigen Synthesis on Virulence and Immunogenicity of Salmonella enterica Serovar Typhimurium
Authors: Kong Qingke , Yang Jiseon , Liu Qing , Alamuri Praveen , Roland Kenneth L. , Curtiss Roy 
Published: July 2011

ABSTRACTLipopolysaccharide (LPS) is a major virulence factor ofSalmonella entericaserovar Typhimurium and is composed of lipid A, core oligosaccharide (C-OS), and O-antigen polysaccharide (O-PS). While the functions of the gene products involved in synthesis of core and O-antigen have been elucidated, the effect of removing O-antigen and core sugars on the virulence and immunogenicity ofSalmonella entericaserovar Typhimurium has not been systematically studied. We introduced nonpolar, defined deletion mutations inwaaG(rfaG),waaI(rfaI),rfaH,waaJ(rfaJ),wbaP(rfbP),waaL(rfaL), orwzy(rfc) into wild-typeS.Typhimurium. The LPS structure was confirmed, and a number ofin vitroandin vivoproperties of each mutant were analyzed. All mutants were significantly attenuated compared to the wild-type parent when administered orally to BALB/c mice and were less invasive in host tissues. Strains with ΔwaaGand ΔwaaImutations, in particular, were deficient in colonization of Peyer's patches and liver. This deficiency could be partially overcome in the ΔwaaImutant when it was administered intranasally. In the context of an attenuated vaccine strain delivering the pneumococcal antigen PspA, all of the mutations tested resulted in reduced immune responses against PspA andSalmonellaantigens. Our results indicate that nonreversible truncation of the outer core is not a viable option for developing a live oralSalmonellavaccine, while awzymutant that retains one O-antigen unit is adequate for stimulating the optimal protective immunity to homologous or heterologous antigens by oral, intranasal, or intraperitoneal routes of administration.
Systematic review of genome-wide expression studies in multiple sclerosis
Authors: Kemppinen A K , Kaprio J , Palotie A , Saarela J 
Published: July 2011

BackgroundAlthough recent genome-wide association studies have identified several genetic variants contributing to the complex aetiology of multiple sclerosis (MS), expression and functional studies are required to further understand its molecular basis.ObjectivesTo identify genes and pathways with differential expression in MS.DesignThe authors conducted a systematic review of seven microarray studies, in which expression in immune cells was compared between MS patients and controls. These studies include a previously unpublished study, which is described here in detail.Results and conclusionAlthough in general the overlap between studies was poor, 229 genes were found to be differentially expressed in MS in at least two studies, of which 11 were in three studies andHSPA1Ain four studies. After excluding the authors' unpublished experiment which may have been affected by certain confounding factors and inclusion of treated subjects, 135 genes were identified in at least two studies. The differentially expressed genes were significantly associated with several immunological pathways, including interleukin (IL)-4, IL-6, IL-17 and glucocorticoid receptor signalling pathways. 15 of the 229 loci have shown some association with MS in published genome-wide association studies (p<0.0001), including three loci with confirmed MS risk variants.