bluuebunny/crossref_metadata_2025_split huggingface dataset:
Multidrug-Resistant
Pseudomonas aeruginosa
Strain That Caused an Outbreak in a Neurosurgery Ward and Its
aac(6′)-Iae
Gene Cassette Encoding a Novel Aminoglycoside Acetyltransferase
Authors: Sekiguchi Jun-ichiro , Asagi Tsukasa , Miyoshi-Akiyama Tohru , Fujino Tomoko , Kobayashi Intetsu , Morita Koji , Kikuchi Yoshihiro , Kuratsuji Tadatoshi , Kirikae Teruo 
Published: August 2005

ABSTRACT

We characterized multidrug-resistant
Pseudomonas aeruginosa
strains isolated from patients involved in an outbreak of catheter-associated urinary tract infections that occurred in a neurosurgery ward of a hospital in Sendai, Japan. Pulsed-field gel electrophoresis of SpeI-, XbaI-, or HpaI-digested genomic DNAs from the isolates revealed that clonal expansion of a
P. aeruginosa
strain designated IMCJ2.S1 had occurred in the ward. This strain possessed broad-spectrum resistance to aminoglycosides, β-lactams, fluoroquinolones, tetracyclines, sulfonamides, and chlorhexidine. Strain IMCJ2.S1 showed a level of resistance to some kinds of disinfectants similar to that of a control strain of
P. aeruginosa
, ATCC 27853. IMCJ2.S1 contained a novel class 1 integron, In113, in the chromosome but not on a plasmid. In113 contains an array of three gene cassettes of
bla
IMP-1
, a novel aminoglycoside resistance gene, and the
aadA1
gene. The aminoglycoside resistance gene, designated
aac(6′)-Iae
, encoded a 183-amino-acid protein that shared 57.1% identity with AAC(6′)-Iq. Recombinant AAC(6′)-Iae protein showed aminoglycoside 6′-
N
-acetyltransferase activity by thin-layer chromatography.
Escherichia coli
expressing exogenous
aac(6′)-Iae
showed resistance to amikacin, dibekacin, isepamicin, kanamycin, netilmicin, sisomicin, and tobramycin but not to arbekacin, gentamicins, or streptomycin. Alterations of
gyrA
and
parC
at the amino acid sequence level were detected in IMCJ2.S1, suggesting that such mutations confer the resistance to fluoroquinolones observed for this strain. These results indicate that
P. aeruginosa
IMCJ2.S1 has developed multidrug resistance by acquiring resistance determinants, including a novel member of the
aac(6′)-I
family and mutations in drug resistance genes.
BEL-1, a Novel Clavulanic Acid-Inhibited Extended-Spectrum β-Lactamase, and the Class 1 Integron In120 in
Pseudomonas aeruginosa
Authors: Poirel Laurent , Brinas Laura , Verlinde Annemie , Ide Louis , Nordmann Patrice 
Published: August 2005

ABSTRACT

Screening by a double-disk synergy test identified a
Pseudomonas aeruginosa
isolate that produced a clavulanic acid-inhibited expanded-spectrum β-lactamase (ESBL). Cloning and sequencing identified a novel ESBL, BEL-1, weakly related to other Ambler class A ESBLs. β-Lactamase BEL-1 hydrolyzed significantly most expanded-spectrum cephalosporins and aztreonam, and its activity was inhibited by clavulanic acid, tazobactam, cefoxitin, moxalactam, and imipenem. This chromosome-encoded ESBL gene was embedded in a class 1 integron containing three other gene cassettes. In addition, this integron was bracketed by Tn
1404
transposon sequences at its right end and by
P. aeruginosa
-specific sequences at its left end.
Short-Course Regimens of Artesunate-Fosmidomycin in Treatment of Uncomplicated
Plasmodium falciparum
Malaria
Authors: Borrmann Steffen , Adegnika Ayola A. , Moussavou Félicien , Oyakhirome Sunny , Esser Gilbert , Matsiegui Pierre-Blaise , Ramharter Michael , Lundgren Ingrid , Kombila Maryvonne , Issifou Saadou , Hutchinson David , Wiesner Jochen , Jomaa Hassan , Kremsner Peter G. 
Published: August 2005

ABSTRACT

Fosmidomycin is effective against malaria, but it needs to be given for ≥4 days when used alone. We conducted a study of 50 children with
Plasmodium falciparum
malaria to evaluate the safety and efficacy of consecutively shortened regimens of artesunate-fosmidomycin (1 to 2 mg/kg of body weight and 30 mg/kg of body weight, respectively; doses given every 12 hours). All dosing regimens were well tolerated. Artesunate-fosmidomycin acted rapidly, resulting in consolidated geometric mean parasite and fever clearance times of 24 h and 15 h, respectively. Treatment regimens of ≥2 days led to cure ratios of 100% by day 14 (39/39; 95% confidence interval [95% CI], 91% to 100%). Most importantly, the 3-day regimen achieved 100% cure on day 28 (10/10; 95% CI, 69% to 100%). Treatment with artesunate-fosmidomycin was associated with transient grade I or II neutropenia (absolute neutrophil counts of 750 to 1,200/μl and 400 to 749/μl, respectively) in six or two patients, respectively. Artesunate-fosmidomycin demonstrates the feasibility and potential value of short-course artemisinin-based combination chemotherapy with rapidly eliminated combination partners.
Analysis of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Subunit Structure/Function in the Context of Infectious Virions and Human Target Cells
Authors: Mulky Alok , Kappes John C. 
Published: August 2005

ABSTRACT

The reverse transcriptase (RT) of all retroviruses is required for synthesis of the viral DNA genome. The human immunodeficiency virus type 1 (HIV-1) RT exists as a heterodimer made up of 51-kDa and 66-kDa subunits. The crystal structure and in vitro biochemical analyses indicate that the p66 subunit of RT is primarily responsible for the enzyme's polymerase and RNase H activities. Since both the p51 and p66 subunits are generated from the same coding region, as part of the Pr160
Gag-Pol
precursor protein, there are inherent limitations for studying subunit-specific function with intact provirus in a virologically relevant context. Our lab has recently described a novel system for studying the RT heterodimer (p51/p66) wherein a LTR-vpr-p51-IRES-p66 expression cassette provided in
trans
to an RT-deleted HIV-1 genome allows precise molecular analysis of the RT heterodimer. In this report, we describe in detail the specific approaches, alternative strategies, and pitfalls that may affect the application of this novel assay for analyzing RT subunit structure/function in infectious virions and human target cells. The ability to study HIV-1 RT subunit structure/function in a physiologically relevant context will advance our understanding of both RT and the process of reverse transcription. The study of antiretroviral drugs in a subunit-specific virologic context should provide new insights into drug resistance and viral fitness. Finally, we anticipate that this approach will help elucidate determinants that mediate p51-p66 subunit interactions, which is essential for structure-based drug design targeting RT heterodimerization.
Quantification of Different Human Alpha Interferon Subtypes and Pegylated Interferon Activities by Measuring MxA Promoter Activation
Authors: François Catherine , Bernard Isabelle , Castelain Sandrine , Charleston Bryan , Fray Martin D. , Capiod Jean-Claude , Duverlie Gilles 
Published: August 2005

ABSTRACT
Alpha interferons (α-IFNs) are potent biologically active proteins synthesized and secreted by somatic cells during viral infection. Quantification of α-IFN concentrations in biological samples is used for diagnosis. More recently, recombinant IFNs have been used as antiviral, antiproliferative, and immunomodulatory therapeutic agents, and particularly for the treatment of chronic hepatitis C virus infection. For this purpose, IFN has recently been coupled to polyethylene glycol (PEG) to improve the pharmacokinetic properties. The measure of α-IFN in biological samples from treated patients could be useful to ensure compliance to therapy and the true IFN activity in relation to viral decay during follow-up. In particular, it could be used to monitor the PEG-IFN concentration in patients treated for hepatitis C virus infection. The most frequently used test is a bioassay based on the antiviral property of the IFN, but the assay is not highly reproducible. Here, we present a reporter test based on MxA promoter activation of chloramphenicol acetyltransferase expression (Mx-CAT). MxA is an antiviral protein induced and tightly regulated by α-IFN. The Mx-CAT assay showed good reproducibility of 15% and was suitable to quantify PEG-IFN and numerous other α-IFN subtypes as well, despite a differential MxA promoter activation in relation with the subtype. A good correlation was obtained with the reporter assay and a commercial enzyme-linked immunosorbent assay on samples from treated patients. This test could be useful for monitoring IFN therapy of chronically infected hepatitis C virus-infected patients treated with the standard IFN, PEG-IFN, and probably forthcoming recombinant IFNs.
Use of
Leishmania donovani
Field Isolates Expressing the Luciferase Reporter Gene in In Vitro Drug Screening
Authors: Ashutosh  , Gupta Suman , Ramesh  , Sundar Shyam , Goyal Neena 
Published: August 2005

ABSTRACT

Currently available primary screens for the selection of candidate antileishmanial compounds are not ideal. These techniques are time-consuming, laborious, and difficult to scale and require macrophages, which limit their use for high-throughput screening. We have developed
Leishmania donovani
field isolates that constitutively express the firefly luciferase reporter gene (
luc
) as a part of an episomal vector. An excellent correlation between parasite number and luciferase activity was observed.
luc
expression was stable, even in the absence of drug selection, for 4 weeks. The transfectants were infective to macrophages, and intracellular amastigotes exhibited luciferase activity. The suitability of these recombinant field isolates for in vitro screening of antileishmanial drugs was established. The luciferase-expressing sodium stibogluconate-resistant cell lines offer a model for the screening of compounds for resistance. The system is in routine use at the Central Drug Research Institute, Lucknow, India, for high-throughput screening of newly synthesized compounds.
Assessment of Effective Renal Plasma Flow, Enzymuria, and Cytokine Release in Healthy Volunteers Receiving a Single Dose of Amphotericin B Desoxycholate
Authors: Pai Manjunath P. , Norenberg Jeffrey P. , Telepak Robert A. , Sidney David S. , Yang Shu 
Published: August 2005

ABSTRACTThe present study assessed potential subclinical markers of amphotericin B (AmB)-related nephrotoxicity and infusion-related reactions (IRR). Subjects were pretreated with diphenhydramine and acetaminophen and received a 500-ml bolus infusion of 0.9% sodium chloride prior to each effective renal plasma flow (ERPF) assessment. ERPF was measured before and after administration of a single 0.25-mg/kg intravenous AmB dose using technetium-99m mercaptoacetyltriglycine. Blood was collected before and 3 h after AmB infusion for tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β) plasma concentrations. Overnight 12-h urine collections were performed before administration of AmB and for 2 nights after administration of AmB and analyzed for α and π glutathione-S-transferases (GSTα and GSTπ, respectively) andN-acetyl-β-d-glucosaminidase (NAG). Six men and six women with mean ± standard deviation (SD) ages of 24.8 ± 5.3 and 28.0 ± 8.5 years, respectively, were studied. Baseline serum creatinine values were within the normal range and were unaltered after administration of AmB. The mean ± SD decrease in ERPF after administration of AmB was significant (P< 0.05) in males (15.7 ± 8.1%) but not females (9.5 ± 14.0%). The GSTπ and GSTα indices increased significantly (P< 0.05) by two to fourfold and returned to baseline in males but were unaltered in females. NAG indices were unaffected by AmB. Six patients experienced an IRR that was associated with increased TNF-α (P< 0.05) but not IL-1β (P= 0.09). These results suggest a potential sex-related difference in AmB-induced nephrotoxicity and provide a rationale for use of ERPF, urine GST, and TNF-α as subclinical markers of polyene-induced toxicity.
Role of
embB
Codon 306 Mutations in
Mycobacterium tuberculosis
Revisited: a Novel Association with Broad Drug Resistance and IS
6110
Clustering Rather than Ethambutol Resistance
Authors: Hazbón Manzour Hernando , Bobadilla del Valle Miriam , Guerrero Marta Inírida , Varma-Basil Mandira , Filliol Ingrid , Cavatore Magali , Colangeli Roberto , Safi Hassan , Billman-Jacobe Helen , Lavender Caroline , Fyfe Janet , García-García Lourdes , Davidow Amy , Brimacombe Michael , León Clara Inés , Porras Tania , Bose Mridula , Chaves Fernando , Eisenach Kathleen D. , Sifuentes-Osornio José , Ponce de León Alfredo , Cave M. Donald , Alland David 
Published: August 2005

ABSTRACT

Mutations at position 306 of
embB
(
embB
306) have been proposed as a marker for ethambutol resistance in
Mycobacterium tuberculosis
; however, recent reports of
embB
306 mutations in ethambutol-susceptible isolates caused us to question the biological role of this mutation. We tested 1,020 clinical
M. tuberculosis
isolates with different drug susceptibility patterns and of different geographical origins for associations between
embB
306 mutations, drug resistance patterns, and major genetic group. One hundred isolates (10%) contained a mutation in
embB
306; however, only 55 of these mutants were ethambutol resistant. Mutations in
embB
306 could not be uniquely associated with any particular type of drug resistance and were found in all three major genetic groups. A striking association was observed between these mutations and resistance to any drug (
P
< 0.001), and the association between
embB
306 mutations and resistance to increasing numbers of drugs was highly significant (
P
< 0.001 for trend). We examined the association between
embB
306 mutations and IS
6110
clustering (as a proxy for transmission) among all drug-resistant isolates. Mutations in
embB
306 were significantly associated with clustering by univariate analysis (odds ratio, 2.44;
P
= 0.004). In a multivariate model that also included mutations in
katG
315,
katG
463,
gyrA
95, and
kasA
269, only mutations in
embB
306 (odds ratio, 2.14;
P
= 0.008) and
katG
315 (odds ratio, 1.99;
P
= 0.015) were found to be independently associated with clustering. In conclusion,
embB
306 mutations do not cause classical ethambutol resistance but may predispose
M. tuberculosis
isolates to the development of resistance to increasing numbers of antibiotics and may increase the ability of drug-resistant isolates to be transmitted between subjects.
Mycobacterium smegmatis
Erm(38) Is a Reluctant Dimethyltransferase
Authors: Madsen Christian Toft , Jakobsen Lene , Douthwaite Stephen 
Published: August 2005

ABSTRACT

The waxy cell walls of mycobacteria provide intrinsic tolerance to a broad range of antibiotics, and this effect is augmented by specific resistance determinants. The inducible determinant
erm
(38) in the nontuberculous species
Mycobacterium smegmatis
confers high resistance to lincosamides and some macrolides, without increasing resistance to streptogramin B antibiotics. This is an uncharacteristic resistance pattern falling between the type I and type II macrolide, lincosamide, and streptogramin B (MLS
B
) phenotypes that are conferred, respectively, by Erm monomethyltransferases and dimethyltransferases. Erm dimethyltransferases are typically found in pathogenic bacteria and confer resistance to all MLS
B
drugs by addition of two methyl groups to nucleotide A2058 in 23S rRNA. We show here by mass spectrometry analysis of the mycobacterial rRNA that Erm(38) is indeed an A2058-specific dimethyltransferase. The activity of Erm(38) is lethargic, however, and only a meager proportion of the rRNA molecules become dimethylated in
M. smegmatis
, while most of the rRNAs are either monomethylated or remain unmethylated. The methylation pattern produced by Erm(38) clarifies the phenotype of
M. smegmatis
, as it is adequate to confer resistance to lincosamides and 14-member ring macrolides such as erythromycin, but it is insufficient to raise the level of resistance to streptogramin B drugs above the already high intrinsic tolerance displayed by this species.
Accumulation of Mutations in both
gyrB
and
parE
Genes Is Associated with High-Level Resistance to Novobiocin in
Staphylococcus aureus
Authors: Fujimoto-Nakamura Mika , Ito Hideaki , Oyamada Yoshihiro , Nishino Takeshi , Yamagishi Jun-ichi 
Published: August 2005

ABSTRACT

Coumarin-resistant mutants of
Staphylococcus aureus
were isolated by three-step selection with novobiocin at different concentrations. Sequencing analysis of the
gyrB
and
parE
genes of the first-, second-, and third-step mutants revealed that successive point mutations first occurred specifically in the
gyrB
gene, followed by a point mutation in the
parE
gene and then an additional point mutation in the
gyrB
gene. These findings demonstrate that DNA gyrase is the primary target and that topoisomerase IV is the secondary target for novobiocin and that the accumulation of point mutations in both the
gyrB
and the
parE
genes is associated with high-level resistance to novobiocin in
S. aureus
. Moreover, our results show that the amino acid substitutions (Asp-89 to Gly and Ser-128 to Leu) found in GyrB are associated with resistance to novobiocin but not to coumermycin A1, suggesting that the interactions of novobiocin and coumermycin A1 with GyrB differ at the molecular level.
Molecular Basis for Increased Susceptibility of Isolates with Atazanavir Resistance-Conferring Substitution I50L to Other Protease Inhibitors
Authors: Yanchunas Joseph , Langley David R. , Tao Li , Rose Ronald E. , Friborg Jacques , Colonno Richard J. , Doyle Michael L. 
Published: August 2005

ABSTRACT
Protease inhibitors (PIs) are highly effective drugs against the human immunodeficiency virus (HIV), yet long-term therapeutic use is limited by emergence of HIV type 1 (HIV-1) protease substitutions that confer cross-resistance to multiple protease inhibitor drugs. Atazanavir is a highly potent HIV protease inhibitor with a distinct resistance profile that includes effectiveness against most HIV-1 isolates resistant to one or two PIs. The signature resistance substitution for atazanavir is I50L, and it is frequently (53%) accompanied by a compensatory A71V substitution that helps restore viability and increases atazanavir resistance levels. We measured the binding affinities of wild-type (WT) and I50L/A71V HIV-1 proteases to atazanavir and other currently approved PIs (ritonavir, lopinavir, saquinavir, nelfinavir, indinavir, and amprenavir) by isothermal titration calorimetry. Remarkably, we find that all of the PIs have 2- to 10-fold increased affinities for I50L/A71V protease, except for atazanavir. The results are also manifested by thermal stability measures of affinity for WT and I50L/A71V proteases. Additional biophysical and enzyme kinetics experiments show I50L/A71V protease is a stable enzyme with catalytic activity that is slightly reduced (34%) relative to the WT. Computational modeling reveals that the unique resistance phenotype of I50L/A71V protease likely originates from bulky tert-butyl groups at P2 and P2′ (specific to atazanavir) that sterically clash with methyl groups on residue L50. The results of this study provide a molecular understanding of the novel hypersusceptibility of atazanavir-resistant I50L/A71V-containing clinical isolates to other currently approved PIs.
High-Throughput Human Immunodeficiency Virus Type 1 (HIV-1) Full Replication Assay That Includes HIV-1 Vif as an Antiviral Target
Authors: Cao Joan , Isaacson Jason , Patick Amy K. , Blair Wade S. 
Published: August 2005

ABSTRACT

Antiviral screens have proved useful for the identification of novel human immunodeficiency virus type 1 (HIV-1) inhibitors. In this study, we describe an HIV-1 full replication (HIV-1 Rep) assay that incorporates all of the targets required for replication in T-cell lines, including the HIV-1 Vif gene. The HIV-1 Rep assay was designed to exhibit optimal sensitivity to late-stage as well as early-stage inhibitors to maximize the likelihood of identification of novel target antiviral compounds in a screen. In addition, the flexibility of the HIV-1 Rep assay allows the rapid evaluation of antiviral compounds against different virus strains in different T-cell lines without significant modification of the assay format. We demonstrate that the HIV-1 Rep assay exhibits characteristics (e.g., a favorable
Z
′ value) compatible with high-throughput screening in a 384-well format. The utility of the HIV-1 Rep assay was demonstrated in a high-throughput screen of >10
6
compounds. To our knowledge, this study represents the first example of an HIV-1 antiviral screen that includes Vif as a functional target and was executed on an industrial scale.
Structure-Function Characterization and Optimization of a Plant-Derived Antibacterial Peptide
Authors: Suarez Mougli , Haenni Marisa , Canarelli Stéphane , Fisch Florian , Chodanowski Pierre , Servis Catherine , Michielin Olivier , Freitag Ruth , Moreillon Philippe , Mermod Nicolas 
Published: August 2005

ABSTRACTCrushed seeds of theMoringa oleiferatree have been used traditionally as natural flocculants to clarify drinking water. We previously showed that one of the seed peptides mediates both the sedimentation of suspended particles such as bacterial cells and a direct bactericidal activity, raising the possibility that the two activities might be related. In this study, the conformational modeling of the peptide was coupled to a functional analysis of synthetic derivatives. This indicated that partly overlapping structural determinants mediate the sedimentation and antibacterial activities. Sedimentation requires a positively charged, glutamine-rich portion of the peptide that aggregates bacterial cells. The bactericidal activity was localized to a sequence prone to form a helix-loop-helix structural motif. Amino acid substitution showed that the bactericidal activity requires hydrophobic proline residues within the protruding loop. Vital dye staining indicated that treatment with peptides containing this motif results in bacterial membrane damage. Assembly of multiple copies of this structural motif into a branched peptide enhanced antibacterial activity, since low concentrations effectively kill bacteria such asPseudomonas aeruginosaandStreptococcus pyogeneswithout displaying a toxic effect on human red blood cells. This study thus identifies a synthetic peptide with potent antibacterial activity against specific human pathogens. It also suggests partly distinct molecular mechanisms for each activity. Sedimentation may result from coupled flocculation and coagulation effects, while the bactericidal activity would require bacterial membrane destabilization by a hydrophobic loop.