bluuebunny/crossref_metadata_2025_split huggingface dataset:
Penicillin-Binding Protein 2 Is Essential for Expression of High-Level Vancomycin Resistance and Cell Wall Synthesis in Vancomycin- Resistant
Staphylococcus aureus
Carrying the Enterococcal
vanA
Gene Complex
Authors: Severin Anatoly , Wu Shang Wei , Tabei Keiko , Tomasz Alexander 
Published: November 2004

ABSTRACT

A combination of biochemical and genetic experiments were performed in order to better understand the mechanism of expression of high-level vancomycin resistance in
Staphylococcus aureus
. The transcription of
pbp2
of the highly vancomycin- and oxacillin-resistant strain COLVA
200
and its mutant derivative with inactivated
mecA
were put under the control of an inducible promoter, and the dependence of oxacillin and vancomycin resistance and cell wall composition on the concentration of the isopropyl-β-
d
-thiogalactopyranoside inducer was determined. The results indicate that
mecA—
the genetic determinant of oxacillin resistance—while essential for oxacillin resistance, is not involved with the expression of vancomycin resistance. Penicillin binding protein 2A, the protein product of
mecA
, appears to be unable to utilize the depsipeptide cell wall precursor produced in the vancomycin-resistant cells for transpeptidation. The key penicillin binding protein essential for vancomycin resistance and for the synthesis of the abnormally structured cell walls characteristic of vancomycin-resistant
S. aureus
(A. Severin, K. Tabei, F. Tenover, M. Chung, N. Clarke, and A. Tomasz, J. Biol. Chem.
279
:3398-3407, 2004) is penicillin binding protein 2.
TMC125, a Novel Next-Generation Nonnucleoside Reverse Transcriptase Inhibitor Active against Nonnucleoside Reverse Transcriptase Inhibitor-Resistant Human Immunodeficiency Virus Type 1
Authors: Andries Koen , Azijn Hilde , Thielemans Theo , Ludovici Donald , Kukla Michael , Heeres Jan , Janssen Paul , De Corte Bart , Vingerhoets Johan , Pauwels Rudi , de Béthune Marie-Pierre 
Published: November 2004

ABSTRACT

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are potent inhibitors of human immunodeficiency virus type 1 (HIV-1); however, currently marketed NNRTIs rapidly select resistant virus, and cross-resistance within the class is extensive. A parallel screening strategy was applied to test candidates from a series of diarylpyrimidines against wild-type and resistant HIV strains carrying clinically relevant mutations. Serum protein binding and metabolic stability were addressed early in the selection process. The emerging clinical candidate, TMC125, was highly active against wild-type HIV-1 (50% effective concentration [EC
50
] = 1.4 to 4.8 nM) and showed some activity against HIV-2 (EC
50
= 3.5 μM). TMC125 also inhibited a series of HIV-1 group M subtypes and circulating recombinant forms and a group O virus. Incubation of TMC125 with human liver microsomal fractions suggested good metabolic stability (15% decrease in drug concentration and 7% decrease in antiviral activity after 120 min). Although TMC125 is highly protein bound, its antiviral effect was not reduced by the presence of 45 mg of human serum albumin/ml, 1 mg of α
1
-acid glycoprotein/ml, or 50% human serum. In an initial screen for activity against a panel of 25 viruses carrying single and double reverse transcriptase amino acid substitutions associated with NNRTI resistance, the EC
50
of TMC125 was <5 nM for 19 viruses, including the double mutants K101E+K103N and K103N+Y181C. TMC125 also retained activity (EC
50
< 100 nM) against 97% of 1,081 recent clinically derived recombinant viruses resistant to at least one of the currently marketed NNRTIs. TMC125 is a potent next generation NNRTI, with the potential for use in individuals infected with NNRTI-resistant virus.
Clinically Relevant Interpretation of Genotype and Relationship to Plasma Drug Concentrations for Resistance to Saquinavir-Ritonavir in Human Immunodeficiency Virus Type 1 Protease Inhibitor-Experienced Patients
Authors: Marcelin Anne-Geneviève , Dalban Cécile , Peytavin Gilles , Lamotte Claire , Agher Rachid , Delaugerre Constance , Wirden Marc , Conan Françoise , Dantin Sylvie , Katlama Christine , Costagliola Dominique , Calvez Vincent 
Published: November 2004

ABSTRACT

It has been shown that virological protease inhibitor (PI) resistance mutations present at the initiation of saquinavir (SQV) plus ritonavir (RTV) therapy in PI-experienced patients are the strongest predictors of virological response. But most of the current resistance algorithms are adapted for unboosted SQV regimens. We applied a stepwise methodology for the development and validation of a clinically relevant genotypic resistance score for an SQV (800 mg twice per day [b.i.d.]) plus RTV (100 mg b.i.d.)-containing regimen. PI-experienced patients treated by this regimen achieved a human immunodeficiency virus plasma viral load (VL) of <200 copies/ml at months 3 to 5 for 41.7% of subjects. Adjusted in a multivariate analysis, taking into account all the confounding factors, such as the nucleoside used, five mutations were combined in a resistance score associated with a reduced virological response to an SQV-plus-RTV regimen: L24I, I62V, V82A/F/T/S, I84V, and L90IM. Patients with isolates harboring 0 to 1 mutation among the score achieved −2.20 log
10
and −1.23 log
10
copies/ml of VL reduction, respectively, while it was −0.27 log
10
copies/ml for those with at least two mutations, classifying the isolates as “no evidence of resistance” (0 or 1 mutation) or “resistance ” (≥2 mutations). The minimum concentration in plasma (
C
min
) of SQV alone was not associated with the virological response. However, the combination of the SQV
C
min
and the genotypic score, expressed as the genotypic inhibitory quotient, was predictive of the virological response, suggesting that the interpretation of SQV concentrations in plasma should be done only in the context of the resistance index provided by viral genotype for PI-experienced patients.
Chalcomycin Biosynthesis Gene Cluster from
Streptomyces bikiniensis
: Novel Features of an Unusual Ketolide Produced through Expression of the
chm
Polyketide Synthase in
Streptomyces fradiae
Authors: Ward Shannon L. , Hu Zhihao , Schirmer Andreas , Reid Ralph , Revill W. Peter , Reeves Christopher D. , Petrakovsky Oleg V. , Dong Steven D. , Katz Leonard 
Published: November 2004

ABSTRACT

Chalcomycin, a 16-membered macrolide antibiotic made by the bacterium
Streptomyces bikiniensis
, contains a 2,3-
trans
double bond and the neutral sugar
d
-chalcose in place of the amino sugar mycaminose found in most other 16-membered macrolides. Degenerate polyketide synthase (PKS)-specific primers were used to amplify DNA fragments from
S. bikiniensis
with very high identity to a unique ketosynthase domain of the tylosin PKS. The resulting amplimers were used to identify two overlapping cosmids encompassing the
chm
PKS. Sequencing revealed a contiguous segment of >60 kb carrying 25 putative genes for biosynthesis of the polyketide backbone, the two deoxysugars, and enzymes involved in modification of precursors of chalcomycin or resistance to it. The
chm
PKS lacks the ketoreductase and dehydratase domains in the seventh module expected to produce the 2,3-double bond in chalcomycin. Expression of PKS in the heterologous host
Streptomyces fradiae
, from which the
tyl
genes encoding the PKS had been removed, resulted in production of at least one novel compound, characterized as a 3-keto 16-membered macrolactone in equilibrium with its 3-
trans
enol tautomer and containing the sugar mycaminose at the C-5 position, in agreement with the structure predicted on the basis of the domain organization of the
chm
PKS. The production of a 3-keto macrolide from the
chm
PKS indicates that a discrete set of enzymes is responsible for the introduction of the 2,3-
trans
double bond in chalcomycin. From comparisons of the open reading frames to sequences in databases, a pathway for the synthesis of nucleoside diphosphate-
d
-chalcose was proposed.
Biochemical Characterization of the THIN-B Metallo-β-Lactamase of
Janthinobacterium lividum
Authors: Docquier Jean-Denis , Lopizzo Teresa , Liberatori Sabrina , Prenna Manuela , Thaller Maria Cristina , Frère Jean-Marie , Rossolini Gian Maria 
Published: November 2004

ABSTRACT

The THIN-B metallo-β-lactamase, a subclass B3 enzyme produced by the environmental species
Janthinobacterium lividum
, was overproduced in
Escherichia coli
by means of a T7-based expression system. The enzyme was purified (>95%) by two ion-exchange chromatography steps and subjected to biochemical analysis. The native THIN-B enzyme is a monomeric protein of 31 kDa. It exhibits the highest catalytic efficiencies with carbapenem substrates and cephalosporins, except for cephaloridine, which acts as a poor inactivator. Individual rate constants for inactivation by chelators were measured, suggesting that inactivation occurred by a mechanism involving formation of a ternary complex.
Molecular and Kinetic Comparison of the Novel Extended-Spectrum β-Lactamases CTX-M-25 and CTX-M-26
Authors: Munday Craig J. , Boyd David A. , Brenwald Nigel , Miller Mark , Andrews Jennifer M. , Wise Richard , Mulvey Michael R. , Hawkey Peter M. 
Published: November 2004

ABSTRACT

CTX-M-25 is a novel extended-spectrum β-lactamase isolated from a single Canadian
Escherichia coli
isolate. Susceptibility testing demonstrated that this enzyme confers resistance to both cefotaxime and ceftazidime, but the level of resistance was reduced with the addition of β-lactamase inhibitors. The
bla
CTX-M-25
gene was detected on a 111-kb plasmid. It is a member of the CTX-M-8 group and has the closest amino acid identity (99%; three amino acid substitutions) with CTX-M-26. The
bla
CTX-M-26
gene was detected on a 100-kb plasmid isolated from a
Klebsiella pneumoniae
strain from the United Kingdom, and plasmid profiling revealed that it showed some homology to the
bla
CTX-M-25
-harboring plasmid. Both CTX-M genes were located downstream of IS
Ecp1
, although the copy upstream of
bla
CTX-M-25
was disrupted by IS
50-A
. Comparative kinetic studies of recombinant CTX-M-25 and CTX-M-26 enzymes showed that CTX-M-25 has a higher level of ceftazidime hydrolysis (
k
cat
values, 33 and 0.005 s
−1
for CTX-M-25 and CTX-M-26, respectively).
Neuraminidase Inhibitor-Rimantadine Combinations Exert Additive and Synergistic Anti-Influenza Virus Effects in MDCK Cells
Authors: Govorkova Elena A. , Fang Hong-Bin , Tan Ming , Webster Robert G. 
Published: November 2004

ABSTRACT
There is insufficient information about combination therapy with approved anti-influenza agents. We tested combinations that paired a neuraminidase (NA) inhibitor (zanamivir, oseltamivir carboxylate, or peramivir) with rimantadine against infection of MDCK cells with H1N1 and H3N2 subtypes of influenza A virus and characterized their mode of interaction. When reduction of extracellular virus was analyzed by individual regression models and three-dimensional representations of the data, all three combinations showed additive and synergistic effects with no cytotoxicity. Maximum synergy against A/New Caledonia/20/99 (H1N1) virus infection was observed with <2.5 μM rimantadine paired with low concentrations of NA inhibitors. All combinations reduced the extracellular yield of A/Panama/2007/99 (H3N2) influenza virus synergistically. However, our findings were different for the cell-associated virus yield. At some drug concentrations, the yield of cell-associated virus was inhibited antagonistically. Therefore, the method of analysis can be a crucial factor in evaluating the interactions of drugs with different mechanisms. We hypothesize that assays based on cell-associated virus yield may underestimate the efficacies of drug combinations that include an NA inhibitor. Taken together, our results suggest that regimens that combine NA inhibitors and rimantadine exert synergistic anti-influenza effects in vitro. These findings provide baseline information for therapeutic testing of the drug combinations in vivo.
Polymorphism in
Plasmodium falciparum
Drug Transporter Proteins and Reversal of In Vitro Chloroquine Resistance by a 9,10-Dihydroethanoanthracene Derivative
Authors: Millet Julie , Alibert Sandrine , Torrentino-Madamet Marylin , Rogier Christophe , Santelli-Rouvier Christiane , Bigot Patricia , Mosnier Joel , Baret Eric , Barbe Jacques , Parzy Daniel , Pradines Bruno 
Published: November 2004

ABSTRACT

BG958 reverses resistance in chloroquine-resistant isolates from different countries. Five mutations in the
Plasmodium falciparum crt
(
pfcrt
) gene resulting in the amino acid changes K76T, M74I, N75E, A220S, and R371I are systematically identified in resistance-reversed Asian, African, and Brazilian parasites which possess the
pfcrt
(CIET) haplotype. In combination with BG958, the activity of chloroquine is increased in parasites with the N86Y mutation in
pfmdr1
.
Glucocorticoids Increase In Vitro and In Vivo Activities of Antibiotics against
Chlamydophila pneumoniae
Authors: Caronzolo Dario , Lucini Valeria , Pannacci Marilou , Grosso Silvia , Colleoni Federica , Fraschini Franco , Scaglione Francesco 
Published: November 2004

ABSTRACT

The in vitro and in vivo antichlamydial activities of dexamethasone and beclomethasone alone and in combination with an antibiotic were tested. In vitro, dexamethasone and beclomethasone decreased the number of inclusion-forming units versus the control number (
P
< 0.001). The combination of glucocorticoids with azithromycin, telithromycin, or levofloxacin was more active than antibiotics used alone (
P
< 0.001). The combination, tested in a murine
Chlamydophila pneumoniae
infection model, produced similar results.
Effects of a Number of Classes of 50S Inhibitors on Stop Codon Readthrough during Protein Synthesis
Authors: Thompson Jill , Pratt Catherine A. , Dahlberg Albert E. 
Published: November 2004

ABSTRACT

The effect of a number of antibiotics on stop codon readthrough during protein synthesis in
Escherichia coli
was examined. Inhibitors which bind close to the entrance of the peptide exit tunnel on the 50S ribosomal subunit promote substantial levels of readthrough, presumably by disrupting the mechanism of peptide release.
Chromosome-Encoded CTX-M-3 from
Kluyvera ascorbata
: a Possible Origin of Plasmid-Borne CTX-M-1-Derived Cefotaximases
Authors: Rodríguez María Margarita , Power Pablo , Radice Marcela , Vay Carlos , Famiglietti Angela , Galleni Moreno , Ayala Juan A. , Gutkind Gabriel 
Published: November 2004

ABSTRACT

A gene identical to plasmid-borne
bla
CTX-M-3
is present in the chromosome of one
Kluyvera ascorbata
strain. It is associated with a structure including an inverted repeat right and an open reading frame 477-like gene probably involved in the mobilization of
bla
CTX-M-3
. Two other
K. ascorbata
strains rendered the previously described
bla
KLUA-9
gene.
A Subinhibitory Concentration of Clarithromycin Inhibits
Mycobacterium avium
Biofilm Formation
Authors: Carter George , Young Lowell S. , Bermudez Luiz E. 
Published: November 2004

ABSTRACT

Mycobacterium avium
causes disseminated infection in immunosuppressed individuals and lung infection in patients with chronic lung diseases.
M. avium
forms biofilm in the environment and possibly in human airways. Antibiotics with activity against the bacterium could inhibit biofilm formation. Clarithromycin inhibits biofilm formation but has no activity against established biofilm.
In Vitro Interactions between Antifungals and Immunosuppressants against
Aspergillus fumigatus
Isolates from Transplant and Nontransplant Patients
Authors: Steinbach William J. , Singh Nina , Miller Jackie L. , Benjamin Daniel K. , Schell Wiley A. , Heitman Joseph , Perfect John R. 
Published: November 2004

ABSTRACT

We performed in vitro antifungal checkerboard testing on 12
Aspergillus fumigatus
clinical isolates (6 transplant recipients and 6 nontransplant patients) with three antifungal agents (amphotericin B, voriconazole, and caspofungin) and three immunosuppressants (FK506, cyclosporine, and rapamycin). We were not able to detect a difference in calcineurin inhibitor antifungal activity against isolates from transplant recipients and nontransplant patients.
Assembly Protein Precursor (pUL80.5 Homolog) of Simian Cytomegalovirus Is Phosphorylated at a Glycogen Synthase Kinase 3 Site and Its Downstream “Priming” Site: Phosphorylation Affects Interactions of Protein with Itself and with Major Capsid Protein
Authors: Casaday Rebecca J. , Bailey Justin R. , Kalb Suzanne R. , Brignole Edward J. , Loveland Amy N. , Cotter Robert J. , Gibson Wade 
Published: November 2004

ABSTRACT
Capsid assembly among the herpes-group viruses is coordinated by two related scaffolding proteins. In cytomegalovirus (CMV), the main scaffolding constituent is called the assembly protein precursor (pAP). Like its homologs in other herpesviruses, pAP is modified by proteolytic cleavage and phosphorylation. Cleavage is essential for capsid maturation and production of infectious virus, but the role of phosphorylation is undetermined. As a first step in evaluating the significance of this modification, we have identified the specific sites of phosphorylation in the simian CMV pAP. Two were established previously to be adjacent serines (Ser156 and Ser157) in a casein kinase II consensus sequence. The remaining two, identified here as Thr231 and Ser235, are within consensus sequences for glycogen synthase kinase 3 (GSK-3) and mitogen-activated protein kinase, respectively. Consistent with Thr231 being a GSK-3 substrate, its phosphorylation required a downstream “priming” phosphate (i.e., Ser235) and was reduced by a GSK-3-specific inhibitor. Phosphorylation of Ser235 converts pAP to an electrophoretically slower-mobility isoform, pAP*; subsequent phosphorylation of pAP* at Thr231 converts pAP* to a still-slower isoform, pAP**. The mobility shift to pAP* was mimicked by substituting an acidic amino acid for either Thr231 or Ser235, but the shift to pAP** required that both positions be phosphorylated. Glu did not substitute for pSer235 in promoting phosphorylation of Thr231. We suggest that phosphorylation of Thr231 and Ser235 causes charge-driven conformational changes in pAP, and we demonstrate that preventing these modifications alters interactions of pAP with itself and with major capsid protein, suggesting a functional significance.
Insertion of a Classical Nuclear Import Signal into the Matrix Domain of the Rous Sarcoma Virus Gag Protein Interferes with Virus Replication
Authors: Garbitt Rachel A. , Bone Karen R. , Parent Leslie J. 
Published: November 2004

ABSTRACT

The Rous sarcoma virus Gag protein undergoes transient nuclear trafficking during virus assembly. Nuclear import is mediated by a nuclear targeting sequence within the MA domain. To gain insight into the role of nuclear transport in replication, we investigated whether addition of a “classical ” nuclear localization signal (NLS) in Gag would affect virus assembly or infectivity. A bipartite NLS derived from nucleoplasmin was inserted into a region of the MA domain of Gag that is dispensable for budding and infectivity. Gag proteins bearing the nucleoplasmin NLS insertion displayed an assembly defect. Mutant virus particles (RC.V8.NLS) were not infectious, although they were indistinguishable from wild-type virions in Gag, Gag-Pol, Env, and genomic RNA incorporation and Gag protein processing. Unexpectedly, postinfection viral DNA synthesis was also normal, as similar amounts of two-long-terminal-repeat junction molecules were detected for RC.V8.NLS and wild type, suggesting that the replication block occurred after nuclear entry of proviral DNA. Phenotypically revertant viruses arose after continued passage in culture, and sequence analysis revealed that the nucleoplasmin NLS coding sequence was deleted from the
gag
gene. To determine whether the nuclear targeting activity of the nucleoplasmin sequence was responsible for the infectivity defect, two critical basic amino acids in the NLS were altered. This virus (RC.V8.KR/AA) had restored infectivity, and the MA.KR/AA protein showed reduced nuclear localization, comparable to the wild-type MA protein. These data demonstrate that addition of a second NLS, which might direct MA and/or Gag into the nucleus by an alternate import pathway, is not compatible with productive virus infection.
In Vitro Characterization of a Simian Immunodeficiency Virus-Human Immunodeficiency Virus (HIV) Chimera Expressing HIV Type 1 Reverse Transcriptase To Study Antiviral Resistance in Pigtail Macaques
Authors: Ambrose Zandrea , Boltz Valerie , Palmer Sarah , Coffin John M. , Hughes Stephen H. , KewalRamani Vineet N. 
Published: November 2004

ABSTRACT

Antiviral resistance is a significant obstacle in the treatment of human immunodeficiency virus type 1 (HIV-1)-infected individuals. Because nonnucleoside reverse transcriptase inhibitors (NNRTIs) specifically target HIV-1 reverse transcriptase (RT) and do not effectively inhibit simian immunodeficiency virus (SIV) RT, the development of animal models to study the evolution of antiviral resistance has been problematic. To facilitate in vivo studies of NNRTI resistance, we examined whether a SIV that causes immunopathogenesis in pigtail macaques could be made sensitive to NNRTIs. Two simian-human immunodeficiency viruses (SHIVs) were derived from the genetic background of SIV
mne
: SIV-RT-YY contains RT substitutions intended to confer NNRTI susceptibility (V181Y and L188Y), and RT-SHIV
mne
contains the entire HIV-1 RT coding region. Both mutant viruses grew to high titers in vitro but had reduced fitness relative to wild-type SIV
mne
. Although the HIV-1 RT was properly processed into p66 and p51 subunits in RT-SHIV
mne
particles, the RT-SHIV
mne
virions had lower levels of RT per viral genomic RNA than HIV-1. Correspondingly, there was decreased RT activity in RT-SHIV
mne
and SIV-RT-YY particles. HIV-1 and RT-SHIV
mne
were similarly susceptible to the NNRTIs efavirenz, nevirapine, and UC781. However, SIV-RT-YY was less sensitive to NNRTIs than HIV-1 or RT-SHIV
mne
. Classical NNRTI resis tance mutations were selected in RT-SHIV
mne
after in vitro drug treatment and were monitored in a sensitive allele-specific real-time RT-PCR assay. Collectively, these results indicate that RT-SHIV
mne
may be a useful model in macaques for the preclinical evaluation of NNRTIs and for studies of the development of drug resistance in vivo.