bluuebunny/crossref_metadata_2025_split huggingface dataset:
Real-Time Reverse Transcription-PCR Quantification of Cytokine mRNA Expression in Golden Syrian Hamster Infected withLeishmania infantumand Treated with a New Amphotericin B Formulation
Authors: Rama Iñiguez S. , Dea-Ayuela M. A. , Sanchez-Brunete J. A. , Torrado J. J. , Alunda J. M. , Bolas-Fernández F. 
Published: March 2006

ABSTRACTA real-time quantitative reverse transcription-PCR assay was developed for the quantification of cytokine mRNA expression in the golden Syrian hamsterMesocricetus auratusinfected withLeishmania infantumand treated with amphotericin B (AMB) formulated in microspheres made of human serum albumin (HSA). Treatment was administered intravenously on days 69, 71, and 73 postinfection (p.i.) with 107metacyclic promastigotes, at doses of 2 and 40 mg/kg of AMB. High infection levels were recorded for untreated animals by day 76 p.i., with parasite loads always about 2 log10per gram higher in the liver than in the spleen. Treatment was highly effective with both doses, but at 40 mg/kg, almost complete parasite elimination was achieved. mRNA expression of gamma interferon (IFN-γ) and, to a lesser extent, tumor necrosis factor alpha (TNF-α) and transforming growth factor beta (TGF-β) in spleen cells was up-regulated in most animals of the untreated group. The mRNA expression of interleukin-4 was strongly down-regulated in untreated as well as treated infected animals. Treatment with the lower dose of AMB-HSA down-regulated the mRNA expression of IFN-γ and TNF-α, with no effect on the deactivating cytokine TGF-β. In contrast, treatment with the higher dose (40 mg/kg) of the formulation caused moderate up-regulation of IFN-γ and TNF-α and strong suppression of TGF-β. Treatment of noninfected animals did not alter the cytokine expression pattern with regard to untreated controls. Our results suggest that treatment ofL. infantum-infected Syrian hamsters with highly effective nontoxic doses of AMB-HSA causes deactivation of the anti-inflammatory cytokine TGF-β, which in turn results in up-regulation of the Th1 cytokines IFN-γ and TNF-α.
Bac 32, a Novel Bacteriocin Widely Disseminated among Clinical Isolates of
Enterococcus faecium
Authors: Inoue Takako , Tomita Haruyoshi , Ike Yasuyoshi 
Published: March 2006

ABSTRACT

A total of 636 vancomycin-resistant
Enterococcus faecium
(VRE) isolates that had been obtained between 1994 and 1999 from the Medical School Hospital of the University of Michigan, Ann Arbor, were tested for bacteriocin production. Two hundred seventy-seven (44%) of the strains were bacteriocinogenic; and 193 of these exhibited activity against
Enterococcus faecium
,
Enterococcus hirae
, and
Enterococcus durans
. Strain VRE200 harbors the highly efficient conjugative gentamicin resistance plasmid pG200 (70 kb) and bacteriocin plasmid pTI1 (12.5 kb). The bacteriocin encoded on pTI1 was designated bacteriocin 32 (Bac 32). Bacteriocin 32 was active against
E. faecium
,
E. hirae
, and
E. durans
but showed no activity against
Listeria monocytogenes
. The Bac 32 genetic locus consists of a bacteriocin gene (
bacA
) and an immunity gene (
bacB
). Neither of these genes showed significant homology to any known bacteriocin determinants. The deduced
bacA
product is 89 amino acids in length, with a putative signal peptide of 19 amino acids at the N terminus. The
bacB
gene encodes a deduced 55-amino-acid protein without a signal sequence. One hundred eighty-nine strains (97.9%) of the 193 strains with activity against the 3 test enterococcal strains gave rise to the expected specific PCR product with a primer specific for
bacA
, indicating that there is a high incidence of Bac 32 production among VRE clinical isolates. Data from Southern analyses of plasmid DNA from 189 of the Bac 32-producing strains with a plasmid pTI1-specific probe suggested that 137 (72.5%) of the strains harbored a pTI1-type plasmid. Bac 32 or Bac 32-type bacteriocin activity and the determinant genes were also identified in 22 (39.3%) of a total of 56 vancomycin-sensitive
E. faecium
clinical isolates, which suggests that this bacteriocin is widely disseminated among
E. faecium
strains.
Gentamicin Causes Apoptosis at Low Concentrations in Renal LLC-PK
1
Cells Subjected to Electroporation
Authors: Servais Hélène , Jossin Yves , Van Bambeke Françoise , Tulkens Paul M. , Mingeot-Leclercq Marie-Paule 
Published: March 2006

ABSTRACT

Gentamicin accumulates in the lysosomes of kidney proximal tubular cells and causes apoptosis at clinically relevant doses. Gentamicin-induced apoptosis can be reproduced with cultured renal cells, but only at high extracellular concentrations (1 to 3 mM; 0.4 to 1.2 g/liter) because of its low level of uptake. We recently showed that gentamicin-induced apoptosis in LLC-PK
1
cells involves a rapid (2-h) permeabilization of lysosomes and activation of the mitochondrial pathway of apoptosis (10 h). We now examine whether the delivery of gentamicin to the cytosol by electroporation would sensitize LLC-PK
1
cells to apoptosis. Cells were subjected to eight pulses (1 ms) at 800 V/cm (square waves) in the presence of gentamicin (3 μM to 3 mM; 1.2 mg/liter to 1.2 g/liter); returned to gentamicin-free medium; and examined at 8 h for their Bax (a marker of mitochondrial pathway activation) contents by Western blotting and competitive reverse transcriptase PCR and at 24 h for apoptosis by 4′,6′-diamidino-2′-phenylindole staining (confirmed by electron microscopy) and for necrosis (by determination of lactate dehydrogenase release). Nonelectroporated cells were incubated with gentamicin for 8 and 24 h. Significant increases in Bax levels (8 h) and apoptosis (24 h) were detected with 0.03 mM (13.2 mg/liter) gentamicin in electroporated cells compared with those achieved with 2 mM (928 mg/liter) in incubated cells. The increase in the Bax level was not associated with an increase in the level of its mRNA but was associated with the accumulation of ubiquitinated forms (probably as a result of impairment of its degradation by the proteasome). Assay of cell-associated gentamicin showed a marked, immediate, but transient accumulation in electroporated cells, whereas a slow, steady uptake was detected in incubated cells. The data indicate that cytosolic gentamicin triggers apoptosis. Sequestration of gentamicin in lysosomes would, to some extent, protect against apoptosis.
Comparative Pharmacokinetics and Pharmacodynamic Target Attainment of Ertapenem in Normal-Weight, Obese, and Extremely Obese Adults
Authors: Chen M. , Nafziger A. N. , Drusano G. L. , Ma L. , Bertino J. S. 
Published: March 2006

ABSTRACT

Little is known of the effects of obesity on ertapenem drug disposition and pharmacodynamics. Thirty healthy volunteers in three body mass index (BMI) groups (10 per group), normal weight (BMI, 18.5 to 24.9 kg/m
2
), class I-II obesity (BMI, 30 to 39.9 kg/m
2
), and class III obesity (BMI, ≥40 kg/m
2
), were administered a 1-g dose of ertapenem. Serum concentrations were obtained over 24 h. Population pharmacokinetic data were obtained using a nonparametric adaptive grid followed by Monte Carlo simulation to determine the probability of obtaining the free drug exposure targets of the time that the free drug concentration remains above the MIC (
f
T
>MIC
) of 20% and 40% for bacteriostatic and maximal bactericidal activity, respectively. Compared to the subjects in the obese groups, area under the concentration-time curve from 0 h to infinity was significantly higher in the normal-weight subjects, whereas the total central compartment volume was higher in the class III obese subjects (
P
≤ 0.05). Achieving a bacteriostatic target of
f
T
>MIC
of 20% with a 90% probability was attained at MICs of ≤0.5 μg/ml for normal-weight subjects. Class I-II and class III obese subjects were able to achieve this target only at a MIC of ≤0.25 μg/ml. For maximal bactericidal activity (
f
T
>MIC
, 40%), no group attained the target at the 90% probability level at any tested MIC. The results suggest that the standard 1-g ertapenem dose may not provide adequate drug exposure for any body mass index classification for MICs in excess of 0.25 to 0.5 μg/ml.
In Vitro Characterization of the Antibacterial Spectrum of Novel Bacterial Type II Topoisomerase Inhibitors of the Aminobenzimidazole Class
Authors: Mani Nagraj , Gross Christian H. , Parsons Jonathan D. , Hanzelka Brian , Müh Ute , Mullin Steve , Liao Yusheng , Grillot Anne-Laure , Stamos Dean , Charifson Paul S. , Grossman Trudy H. 
Published: March 2006

ABSTRACT

Antibiotics with novel mechanisms of action are becoming increasingly important in the battle against bacterial resistance to all currently used classes of antibiotics. Bacterial DNA gyrase and topoisomerase IV (topoIV) are the familiar targets of fluoroquinolone and coumarin antibiotics. Here we present the characterization of two members of a new class of synthetic bacterial topoII ATPase inhibitors: VRT-125853 and VRT-752586. These aminobenzimidazole compounds were potent inhibitors of both DNA gyrase and topoIV and had excellent antibacterial activities against a wide spectrum of problematic pathogens responsible for both nosocomial and community-acquired infections, including staphylococci, streptococci, enterococci, and mycobacteria. Consistent with the novelty of their structures and mechanisms of action, antibacterial potency was unaffected by commonly encountered resistance phenotypes, including fluoroquinolone resistance. In time-kill assays, VRT-125853 and VRT-752586 were bactericidal against
Staphylococcus aureus
,
Streptococcus pneumoniae
,
Enterococcus faecalis
, and
Haemophilus influenzae
, causing 3-log reductions in viable cells within 24 h. Finally, similar to the fluoroquinolones, relatively low frequencies of spontaneous resistance to VRT-125853 and VRT-752586 were found, a property consistent with their in vitro dual-targeting activities.
Nanodisk-Associated Amphotericin B Clears
Leishmania major
Cutaneous Infection in Susceptible BALB/c Mice
Authors: Nelson Keith G. , Bishop Jeanette V. , Ryan Robert O. , Titus Richard 
Published: March 2006

ABSTRACT

Nanometer-scale, apolipoprotein-stabilized phospholipid bilayer disk complexes (nanodisks [ND]) harboring the toxic and poorly soluble antileishmanial agent amphotericin B (AMB) were examined for efficacy in treatment of
Leishmania major
-infected BALB/c mice (
Mus musculus
).
L. major
-infected mice were intraperitoneally (i.p.) treated with AMB-ND in 0-, 1-, and 5-mg/kg doses at 24 h, 48 h, and 4, 7, 14, and 21 days postinfection in two experiments.
L. major
-infected mice were i.p. treated with phosphate-buffered saline, 5 mg/kg AMB-ND, or 5 mg/kg lipid-associated amphotericin B (liposomal amphotericin B, AmBisome) at 24 h, 48 h, and 10, 20, 30, and 40 days postinfection in one experiment. Parasite numbers, footpad lesion size progression, and development of cytokine responses were assayed at days 7, 15, 30, 50, 140, and 250 or at days 14, 30, 50, 95, and 140 postinfection. Mice administered AMB-ND in 1- or 5-mg/kg doses were significantly protected from
L. major
, displaying decreases in lesion size and parasite burden, particularly at the 5-mg/kg dosage level. In contrast to the i.p. treated AmBisome group, BALB/c mice treated with i.p. AMB-ND completely cleared an
L. major
infection by 140 to 250 days postinfection, with no lesions remaining and no parasites isolated from infected animals. Restimulated mixed lymphocyte culture cytokine responses (interleukin-4 [IL-4], IL-12, IL-10, NO, and gamma interferon) were unchanged by AMB-ND administration compared to controls. The marked clearance of
Leishmania
parasites from a susceptible strain of mice without an appreciable change in the cytokine response suggests that AMB-ND represent a potentially useful formulation for treatment of intrahistiocytic organisms.
Rapid Microbiologic and Pharmacologic Evaluation of Experimental Compounds against
Mycobacterium tuberculosis
Authors: Gruppo Veronica , Johnson Christine M. , Marietta Karen S. , Scherman Hataichanok , Zink Erin E. , Crick Dean C. , Adams Linda B. , Orme Ian M. , Lenaerts Anne J. 
Published: March 2006

ABSTRACT

The assessment of physiochemical and pharmacological properties at early stages of drug discovery can accelerate the conversion of hits and leads into candidates for further development. A strategy for streamlined evaluation of compounds against
Mycobacterium tuberculosis
in the early preclinical stage is presented in this report. As a primary assay to rapidly select experimental compounds with sufficient in vitro activity, the growth inhibition microtiter plate assay was devised as an alternative to current methods. This microdilution plate assay is a liquid culture method based on spectrophotometric readings of the bacillary growth. The performance of this method was compared to the performance of two established susceptibility methods using clinical available tuberculosis (TB) drugs. Data generated from all three assays were similar for all of the tested compounds. A second simple bioassay was devised to assess the oral bioavailability of compounds prior to extensive in vivo efficacy testing. The bioassay estimates drug concentrations in collected serum samples by a microdilution MIC plate method using
M. tuberculosis
. In the same assay, the MIC of the compound is also determined in the presence of 10% mouse serum as an indication of protein binding. The method was validated using different clinically available TB drugs, and results are discussed in this report. With these methodological advances, screening of compounds against tuberculosis in the preclinical phase will be rapid, can be adapted to semi-high-throughput screening, and will add relevant physicochemical and basic pharmacological criteria to the decision process of drug discovery.
Activity of Cecropin A-Melittin Hybrid Peptides against Colistin-Resistant Clinical Strains of
Acinetobacter baumannii
: Molecular Basis for the Differential Mechanisms of Action
Authors: Saugar José María , Rodríguez-Hernández María Jesús , de la Torre Beatriz G. , Pachón-Ibañez María Eugenia , Fernández-Reyes María , Andreu David , Pachón Jerónimo , Rivas Luis 
Published: March 2006

ABSTRACT

Acinetobacter baumannii
has successfully developed resistance against all common antibiotics, including colistin (polymyxin E), the last universally active drug against this pathogen. The possible widespread distribution of colistin-resistant
A. baumannii
strains may create an alarming clinical situation. In a previous work, we reported differences in lethal mechanisms between polymyxin B (PXB) and the cecropin A-melittin (CA-M) hybrid peptide CA(1-8)M(1-18) (KWKLFKKIGIGAVLKVLTTGLPALIS-NH
2
) on colistin-susceptible strains (J. M. Saugar, T. Alarcón, S. López-Hernández, M. López-Brea, D. Andreu, and L. Rivas, Antimicrob. Agents Chemother.
46:
875-878, 2002). We now demonstrate that CA(1-8)M(1-18) and three short analogues, namely CA(1-7)M(2-9) (KWKLFKKIGAVLKVL-NH
2
), its N
α
-octanoyl derivative (Oct-KWKLFKKIGAVLKVL-NH
2
), and CA(1-7)M(5-9) (KWKLLKKIGAVLKVL-NH
2
) are active against two colistin-resistant clinical strains. In vitro, resistance to colistin sulfate was targeted to the outer membrane, as spheroplasts were equally lysed by a given peptide, regardless of their respective level of colistin resistance. The CA-M hybrids were more efficient than colistin in displacing lipopolysaccharide-bound dansyl-polymyxin B from colistin-resistant but not from colistin-susceptible strains. Similar improved performance of the CA-M hybrids in permeation of the inner membrane was observed, regardless of the resistance pattern of the strain. These results argue in favor of a possible use of CA-M peptides, and by extension other antimicrobial peptides with similar features, as alternative chemotherapy in colistin-resistant
Acinetobacter
infections.
Clinical and Economic Impact of Bacteremia with Extended- Spectrum-β-Lactamase-Producing
Enterobacteriaceae
Authors: Schwaber Mitchell J. , Navon-Venezia Shiri , Kaye Keith S. , Ben-Ami Ronen , Schwartz David , Carmeli Yehuda 
Published: March 2006

ABSTRACT

We studied outcomes of extended-spectrum β-lactamase (ESBL) production in
Enterobacteriaceae
bacteremia. Inpatients with bacteremia caused by ESBL-producing
Escherichia coli
,
Klebsiella
spp., or
Proteus
spp. (cases) were compared with patients with bacteremia caused by non-ESBL producers (controls). Outcomes included mortality, mortality due to infection, length of stay (LOS), delay in appropriate therapy (DAT), discharge to a chronic care facility, and hospital cost. Ninety-nine cases and 99 controls were enrolled. Thirty-five percent of cases died, versus 18% of controls (odds ratio [OR], 2.5; 95% confidence interval [CI], 1.3 to 4.7;
P
= 0.01). Thirty percent of cases died due to infection, versus 16% of controls (OR, 2.3; 95% CI, 1.1 to 4.5;
P
= 0.03). The median LOS after bacteremia for cases was 11 days (interquartile range, 5 to 21), versus 5 days for controls (interquartile range, 3 to 9) (
P
< 0.001). DAT occurred in 66% of cases, versus 7% of controls (OR, 25.1; 95% CI, 10.5 to 60.2;
P
< 0.001). Cases were more likely than controls to be discharged to chronic care (52% versus 21%; OR, 4.0; 95% CI, 1.9 to 8.3;
P
< 0.001). The average hospital cost for cases was 65,509 Israeli shekels, versus 23,538 shekels for controls (
P
< 0.001). After adjusting for differences between groups by using multivariable analysis, ESBL production remained a significant predictor of mortality (OR, 3.6; 95% CI, 1.4 to 9.5;
P
= 0.008), increased LOS (1.56-fold;
P
= 0.001), DAT (OR, 25.1; 95% CI, 10.5 to 60.2;
P
< 0.001), and increased cost (1.57-fold;
P
= 0.003). The mean increase in equivalent cost attributable to ESBL production was $9,620. ESBL production was associated with severe adverse outcomes, including higher overall and infection-related mortality, increased LOS, DAT, discharge to chronic care, and higher costs.
MepR, a Repressor of the
Staphylococcus aureus
MATE Family Multidrug Efflux Pump MepA, Is a Substrate-Responsive Regulatory Protein
Authors: Kaatz Glenn W. , DeMarco Carmen E. , Seo Susan M. 
Published: March 2006

ABSTRACT

The
mepRAB
gene cluster of
Staphylococcus aureus
encodes a MarR family repressor (MepR; known to repress
mepA
expression), a MATE family multidrug efflux pump (MepA), and a protein of unknown function (MepB). In this report, we show that MepR also is autoregulatory, repressing the expression of its own gene. Exposure of strains containing a
mepR
::
lacZ
fusion with
mepR
provided in
trans
under the control of an inducible promoter, or a
mepA
::
lacZ
fusion alone, to subinhibitory concentrations of MepA substrates resulted in variably increased expression mainly of
mepA
. Mobility shift assays revealed that MepR binds upstream of
mepR
and
mepA
, with an apparently higher affinity for the
mepA
binding site. MepA substrates abrogated MepR binding to each site in a differential manner, with the greatest effect observed on the MepR-
mepA
operator interaction. DNase I footprinting identified precise binding sites which included promoter motifs, inverted repeats, and transcription start sites for
mepR
and
mepA
, as well as a conserved GTTAG motif, which may be a signature recognition sequence for MepR. Analogous to other multidrug efflux pump regulatory proteins such as QacR, the substrate-MepR interaction likely results in its dissociation from its
mepA
, and in a more limited fashion its
mepR
, operator sites and relief of its repressive effect. The enhanced effect of substrates on
mepA
compared to
mepR
expression, and on the MepR-
mepA
operator interaction, results in significant relief of
mepA
and relative maintenance of
mepR
repression, leading to increased MepA protein unimpeded by MepR when the need for detoxification exists.
In Vitro Analysis of IS
Ecp1B
-Mediated Mobilization of Naturally Occurring β-Lactamase Gene
bla
CTX-M
of
Kluyvera ascorbata
Authors: Lartigue Marie-Frédérique , Poirel Laurent , Aubert Daniel , Nordmann Patrice 
Published: March 2006

ABSTRACT

IS
Ecp1B
has been reported to be associated with and to mobilize the emerging expanded-spectrum β-lactamase
bla
CTX-M
genes in
Enterobacteriaceae
. Thus, the ability of this insertion sequence to mobilize the
bla
CTX-M-2
gene was tested from its progenitor,
Kluyvera ascorbata
. Insertions of IS
Ecp1B
upstream of the
bla
CTX-M-2
gene in
K. ascorbata
reference strain CIP7953 were first selected with cefotaxime (0.5 and 2 μg/ml). In those cases, IS
Ecp1B
brought promoter sequences enhancing
bla
CTX-M-2
expression in
K. ascorbata
. Then, IS
Ecp1B
-mediated mobilization of the
bla
CTX-M-2
gene from
K. ascorbata
to
Escherichia coli
J53 was attempted. The transposition frequency of IS
Ecp1B-bla
CTX-M-2
occurred at (6.4 ± 0.5) × 10
−7
in
E. coli
. Cefotaxime, ceftazidime, and piperacillin enhanced transposition, whereas amoxicillin, cefuroxime, and nalidixic acid did not. Transposition was also enhanced when studied at 40°C.
Evaluation of Amphotericin B Interpretive Breakpoints for
Candida
Bloodstream Isolates by Correlation with Therapeutic Outcome
Authors: Park Benjamin J. , Arthington-Skaggs Beth A. , Hajjeh Rana A. , Iqbal Naureen , Ciblak Meral A. , Lee-Yang Wendy , Hairston Mario D. , Phelan Maureen , Plikaytis Brian D. , Sofair Andre N. , Harrison Lee H. , Fridkin Scott K. , Warnock David W. 
Published: March 2006

ABSTRACT

One hundred seven
Candida
bloodstream isolates (51
C. albicans
, 24
C. glabrata
, 13
C. parapsilosis
, 13
C. tropicalis
, 2
C. dubliniensis
, 2
C. krusei
, and 2
C. lusitaniae
strains) from patients treated with amphotericin B alone underwent in vitro susceptibility testing against amphotericin B using five different methods. Fifty-four isolates were from patients who failed treatment, defined as death 7 to 14 days after the incident candidemia episode, having persistent fever of ≥5 days' duration after the date of the incident candidemia, or the recurrence of fever after two consecutive afebrile days while on antifungal treatment. MICs were determined by using the Clinical Laboratory Standards Institute (formally National Committee for Clinical Laboratory Standards) broth microdilution procedure with two media and by using Etest. Minimum fungicidal concentrations (MFCs) were also measured in two media. Broth microdilution tests with RPMI 1640 medium generated a restricted range of MICs (0.125 to 1 μg/ml); the corresponding MFC values ranged from 0.5 to 4 μg/ml. Broth microdilution tests with antibiotic medium 3 produced a broader distribution of MIC and MFC results (0.015 to 0.25 μg/ml and 0.06 to 2 μg/ml, respectively). Etest produced the widest distribution of MICs (0.094 to 2 μg/ml). However, none of the test formats studied generated results that significantly correlated with therapeutic success or failure.