bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Antimalarial Trioxaquine DU1301 Alkylates Heme in Malaria-Infected Mice
Authors: Bousejra-El Garah Fatima , Claparols Catherine , Benoit-Vical Françoise , Meunier Bernard , Robert Anne 
Published: June 2008

ABSTRACT

The in vivo alkylation of heme by the antimalarial trioxaquine DU1301 afforded covalent heme-drug adducts that were detected in the spleens of
Plasmodium
sp.-infected mice. This result indicates that the alkylation capacities of trioxaquines in mammals infected with
Plasmodium
strains are similar to that of artemisinin, a natural antimalarial trioxane-containing drug.
Kinetic Characterization of VIM-7, a Divergent Member of the VIM Metallo-β-Lactamase Family
Authors: Samuelsen Ørjan , Castanheira Mariana , Walsh Timothy R. , Spencer James 
Published: June 2008

ABSTRACT
Purified recombinant VIM-7 possesses efficient penicillinase and carbapenemase activities comparable to those of VIM-2. Cephalosporinase activity was variable and generally lower than those of VIM-1 and VIM-2. A homology model suggests that the VIM-7 Tyr-218 Phe substitution may be responsible for the reduced catalytic efficiency against certain cephalosporins, including ceftazidime and cefepime.
Molecular Mechanism of Hepatitis C Virus Replicon Variants with Reduced Susceptibility to a Benzofuran Inhibitor, HCV-796
Authors: Howe Anita Y. M. , Cheng Huiming , Johann Stephen , Mullen Stanley , Chunduru Srinivas K. , Young Dorothy C. , Bard Joel , Chopra Rajiv , Krishnamurthy Girija , Mansour Tarek , O'Connell John 
Published: June 2008

ABSTRACT

HCV-796 selectively inhibits hepatitis C virus (HCV) NS5B RNA-dependent RNA polymerase. In hepatoma cells containing a genotype 1b HCV replicon, HCV-796 reduced HCV RNA levels by 3 to 4 log
10
HCV copies/μg total RNA (the concentration of the compound that inhibited 50% of the HCV RNA level was 9 nM). Cells bearing replicon variants with reduced susceptibility to HCV-796 were generated in the presence of HCV-796, followed by G418 selection. Sequence analysis of the NS5B gene derived from the replicon variants revealed several amino acid changes within 5 Å of the drug-binding pocket. Specifically, mutations were observed at Leu314, Cys316, Ile363, Ser365, and Met414 of NS5B, which directly interact with HCV-796. The impacts of the amino acid substitutions on viral fitness and drug susceptibility were examined in recombinant replicons and NS5B enzymes with the single-amino-acid mutations. The replicon variants were 10- to 1,000-fold less efficient in forming colonies in cells than the wild-type replicon; the S365L variant failed to establish a stable cell line. Other variants (L314F, I363V, and M414V) had four- to ninefold-lower steady-state HCV RNA levels. Reduced binding affinity with HCV-796 was demonstrated in an enzyme harboring the C316Y mutation. The effects of these resistance mutations were structurally rationalized using X-ray crystallography data. While different levels of resistance to HCV-796 were observed in the replicon and enzyme variants, these variants retained their susceptibilities to pegylated interferon, ribavirin, and other HCV-specific inhibitors. The combined virological, biochemical, biophysical, and structural approaches revealed the mechanism of resistance in the variants selected by the potent polymerase inhibitor HCV-796.
Role of Porins for Uptake of Antibiotics by
Mycobacterium smegmatis
Authors: Danilchanka Olga , Pavlenok Mikhail , Niederweis Michael 
Published: June 2008

ABSTRACT

The outer membrane of mycobacteria presents an effective permeability barrier for many antibiotics. Transport pathways across this membrane are unknown for most drugs. Here, we examined which antibiotics utilize the porin pathway across the outer membrane of the model organism
Mycobacterium smegmatis
. Deletion of the porins MspA and MspC drastically increased the resistance of
M. smegmatis
ML10 to β-lactam antibiotics, while its β-lactamase activity remained unchanged. These results are consistent with the ninefold-reduced outer membrane permeability of the
M. smegmatis
porin mutants for cephaloridine and strongly indicate that β-lactam antibiotics rely on the porin pathway. The porin mutant ML10 accumulated less chloramphenicol and norfloxacin and was less susceptible to these antibiotics than wild-type
M. smegmatis
. These results demonstrated that small and hydrophilic antibiotics use the Msp porins for entering the cell. In contrast to norfloxacin, the hydrophobic moxifloxacin was 32-fold more effective in inhibiting the growth of
M. smegmatis
, presumably because it was able to diffuse through the lipid membrane. Structural models indicated that erythromycin, kanamycin, and vancomycin are too large to move through the MspA channel. This study presents the first experimental evidence that hydrophilic fluoroquinolones and chloramphenicol diffuse through porins in mycobacteria. Thus, mutations resulting in less efficient porins or lower porin expression levels are likely to represent a mechanism for the opportunistic pathogens
M. avium
,
M. chelonae
, and
M. fortuitum
, which have Msp-like porins, to acquire resistance to fluoroquinolones.
Rifampin and Rifaximin Resistance in Clinical Isolates of
Clostridium difficile
Authors: O'Connor Jennifer R. , Galang Minerva A. , Sambol Susan P. , Hecht David W. , Vedantam Gayatri , Gerding Dale N. , Johnson Stuart 
Published: June 2008

ABSTRACT

Rifaximin, a poorly absorbed rifamycin derivative, is a promising alternative for the treatment of
Clostridium difficile
infections. Resistance to this agent has been reported, but no commercial test for rifaximin resistance exists and the molecular basis of this resistance has not been previously studied in
C. difficile
. To evaluate whether the rifampin Etest would be a suitable substitute for rifaximin susceptibility testing in the clinical setting, we analyzed the in vitro rifaximin susceptibilities of 80 clinical isolates from our collection by agar dilution and compared these results to rifampin susceptibility results obtained by agar dilution and Etest. We found rifaximin susceptibility data to agree with rifampin susceptibility; the MICs of both antimicrobials for all isolates were either very low or very high. Fourteen rifaximin-resistant (MIC, ≥32 μg/ml) unique isolates from patients at diverse locations in three countries were identified. Molecular typing analysis showed that nine (64%) of these isolates belonged to the epidemic BI/NAP1/027 group that is responsible for multiple outbreaks and increased disease severity in the United Kingdom, Europe, and North America. The molecular basis of rifaximin and rifampin resistance in these isolates was investigated by sequence analysis of
rpoB
, which encodes the β subunit of RNA polymerase, the target of rifamycins. Resistance-associated
rpoB
sequence differences that resulted in specific amino acid substitutions in an otherwise conserved region of RpoB were found in all resistant isolates. Seven different RpoB amino acid substitutions were identified in the resistant isolates, which were divided into five distinct groups by restriction endonuclease analysis typing. These results suggest that the amino acid substitutions associated with rifamycin resistance were independently derived rather than disseminated from specific rifamycin-resistant clones. We propose that rifaximin resistance in
C. difficile
results from mutations in RpoB and that rifampin resistance predicts rifaximin resistance for this organism.
Evaluation of Etest Method for Determining Isavuconazole MICs against
Cryptococcus gattii
and
Cryptococcus neoformans
Authors: Thompson George R. , Fothergill Annette W. , Wiederhold Nathan P. , Vallor Ana C. , Wickes Brian L. , Patterson Thomas F. 
Published: June 2008

ABSTRACT

We compared Etest with broth microdilution testing for isavuconazole activity against 92
Cryptococcus
isolates. A 97.8% agreement was found between these methods, without major discrepancies (>2-well dilution difference). Our findings support the use of the Etest methodology as a reliable method for the determination of MICs against
Cryptococcus
spp.
First Assessment in Humans of the Safety, Tolerability, Pharmacokinetics, and Ex Vivo Pharmacodynamic Antimalarial Activity of the New Artemisinin Derivative Artemisone
Authors: Nagelschmitz Johannes , Voith Barbara , Wensing Georg , Roemer Axel , Fugmann Burkhard , Haynes Richard K. , Kotecka Barbara M. , Rieckmann Karl H. , Edstein Michael D. 
Published: June 2008

ABSTRACT

In preclinical studies, artemisone (BAY 44-9585), a new artemisinin derivative, was shown to possess enhanced efficacy over artesunate, and it does not possess the neurotoxicity characteristic of the current artemisinins. In a phase I program with double-blind, randomized, placebo-controlled, single and multiple ascending oral-dose studies, we evaluated the safety, tolerability, pharmacokinetics, and ex vivo pharmacodynamic antimalarial activity of artemisone. Single doses (10, 20, 30, 40, and 80 mg) and multiple doses (40 and 80 mg daily for 3 days) of artemisone were administered orally to healthy subjects. Plasma concentrations of artemisone and its metabolites were measured by liquid chromatography/tandem mass spectrometry (LC/MS-MS). Artemisone was well tolerated, with no serious adverse events and no clinically relevant changes in laboratory and vital parameters. The pharmacokinetics of artemisone over the 10- to 80-mg range demonstrated dose linearity. After the single 80-mg dose, artemisone had a geometric mean maximum concentration of 140.2 ng/ml (range, 86.6 to 391.0), a short elimination half-life (
t
1/2
) of 2.79 h (range, 1.56 to 4.88), a high oral clearance of 284.1 liters/h (range, 106.7 to 546.7), and a large volume of distribution of 14.50 liters/kg (range, 3.21 to 51.58). Due to artemisone's short
t
1/2
, its pharmacokinetics were comparable after single and multiple dosing. Plasma samples taken after multiple dosing showed marked ex vivo pharmacodynamic antimalarial activities against two multidrug-resistant
Plasmodium falciparum
lines. Artemisone equivalent concentrations measured by bioassay revealed higher activity than artemisone measured by LC/MS-MS, confirming the presence of active metabolites. Comparable to those of other artemisinin's, artemisone's
t
1/2
is well suited for artemisinin-based combination therapy for the treatment of
P. falciparum
malaria.
Comparative Evaluation of Virus Transmission Inhibition by Dual-Acting Pyrimidinedione Microbicides Using the Microbicide Transmission and Sterilization Assay
Authors: Watson Karen M. , Buckheit Christa E. , Buckheit Robert W. 
Published: June 2008

ABSTRACT
In the absence of a fully effective human immunodeficiency virus (HIV) vaccine, topical microbicides represent an important strategy for preventing the transmission of HIV through sexual intercourse, the predominant mode of HIV transmission worldwide. Although a comprehensive understanding of HIV transmission has not yet emerged in the microbicide field, it is likely the result of rapid infection of monocyte-derived cells in the vaginal mucosa by CCR5-tropic viruses. Inhibition of HIV transmission requires agents that prevent entry, fusion, reverse transcription, or other preintegrative replication events or agents which directly inactivate HIV or modulate the target cells to render them uninfectible. In vitro assays typically used to evaluate the ability of a microbicide to prevent virus transmission use epithelial or human osteosarcoma-derived cells or immune cells more relevant to the development of anti-HIV therapeutic agents and quantify virus production at short time intervals following infection. We have developed a microbicide transmission and sterilization assay (MTSA) to more sensitively and quantitatively evaluate virus transmission in cell culture in the presence of microbicidal compounds. Results obtained with the MTSA demonstrate that the inhibitory capacity of microbicides is often overestimated in short-term transmission inhibition assays, while some compounds yield equivalent inhibitory results, indicating a biological relevance for the MTSA-based evaluations to identify superior potent microbicides. The MTSA defines the concentration of the microbicide required to totally suppress the transmission of virus in cell culture and may thus help define the effective concentration of the microbicide required in a formulated microbicide product.
Inhibition of OXA-1 β-Lactamase by Penems
Authors: Bethel Christopher R. , Distler Anne M. , Ruszczycky Mark W. , Carey Marianne P. , Carey Paul R. , Hujer Andrea M. , Taracila Magda , Helfand Marion S. , Thomson Jodi M. , Kalp Matthew , Anderson Vernon E. , Leonard David A. , Hujer Kristine M. , Abe Takao , Venkatesan Aranapakam M. , Mansour Tarek S. , Bonomo Robert A. 
Published: June 2008

ABSTRACT

The partnering of a β-lactam with a β-lactamase inhibitor is a highly effective strategy that can be used to combat bacterial resistance to β-lactam antibiotics mediated by serine β-lactamases (EC 3.2.5.6). To this end, we tested two novel penem inhibitors against OXA-1, a class D β-lactamase that is resistant to inactivation by tazobactam. The

K
i

of each penem inhibitor for OXA-1 was in the nM range (

K
i

of penem 1, 45 ± 8 nM;

K
i

of penem 2, 12 ± 2 nM). The first-order rate constant for enzyme and inhibitor complex inactivation of penems 1 and 2 for OXA-1 β-lactamase were 0.13 ± 0.01 s
−1
and 0.11 ± 0.01 s
−1
, respectively. By using an inhibitor-to-enzyme ratio of 1:1, 100% inactivation was achieved in ≤900 s and the recovery of OXA-1 β-lactamase activity was not detected at 24 h. Covalent adducts of penems 1 and 2 (changes in molecular masses, +306 ± 3 and +321 ± 3 Da, respectively) were identified by electrospray ionization mass spectrometry (ESI-MS). After tryptic digestion of OXA-1 inactivated by penems 1 and 2, ESI-MS and matrix-assisted laser desorption ionization-time-of-flight MS identified the adducts of 306 ± 3 and 321 ± 3 Da attached to the peptide containing the active-site Ser67. The base hydrolysis of penem 2, monitored by serial
1
H nuclear magnetic resonance analysis, suggested that penem 2 formed a linear imine species that underwent 7-endo-trig cyclization to ultimately form a cyclic enamine, the 1,4-thiazepine derivative. Susceptibility testing demonstrated that the penem inhibitors at 4 mg/liter effectively restored susceptibility to piperacillin. Penem β-lactamase inhibitors which demonstrate high affinities and which form long-lived acyl intermediates may prove to be extremely useful against the broad range of inhibitor-resistant serine β-lactamases present in gram-negative bacteria.
Molecular Basis of UropathogenicEscherichia coliEvasion of the Innate Immune Response in the Bladder
Authors: Billips Benjamin K. , Schaeffer Anthony J. , Klumpp David J. 
Published: June 2008

ABSTRACTIn the urinary tract, the innate immune system detects conserved bacterial components and responds to infection by activating the proinflammatory transcription factor NF-κB, resulting in cytokine secretion and neutrophil recruitment. UropathogenicEscherichia coli(UPEC), however, has been shown to evade the host innate immune response by suppressing NF-κB activation in urothelial cells, which results in decreased cytokine secretion and increased urothelial apoptosis. To understand the molecular basis of UPEC modulation of inflammation, we performed a genetic screen with UPEC strain NU14 to identify genes which are required for modulation of urothelial cytokine secretion. Disruption ofampG(peptidoglycan permease),waaL(lipopolysaccharide O antigen ligase), oralr(alanine racemase) resulted in increased urothelial interleukin-8 (IL-8) and IL-6 release from urothelial cell cultures. Targeted deletion of these genes also resulted in elevated urothelial cytokine production during UPEC infection. Conditioned media from bacterial cultures of NU14 ΔampGand NU14 ΔwaaLcontained a heat-stable factor(s) which stimulated greater urothelial IL-8 secretion than that in NU14-conditioned medium. In a mouse model of urinary tract infection, NU14 ΔampG, NU14 ΔwaaL, and NU14 Δalrwere attenuated compared to wild-type NU14 and showed reduced fitness in competition experiments. Instillation of NU14 ΔampGor NU14 ΔwaaLincreased bladder neutrophil recruitment, indicating that enhanced urothelial cytokine secretion during urinary tract infection results in an altered host response. Thus, UPEC evasion of innate immune detection of bacterial components, such as lipopolysaccharide and peptidoglycan fragments, is likely an important factor in the ability of UPEC to colonize the urinary tract.
EspF Interacts with Nucleation-Promoting Factors To Recruit Junctional Proteins into Pedestals for Pedestal Maturation and Disruption of Paracellular Permeability
Authors: Peralta-Ramírez Janneth , Hernandez J. Manuel , Manning-Cela Rebeca , Luna-Muñoz José , Garcia-Tovar Carlos , Nougayréde Jean-Philippe , Oswald Eric , Navarro-Garcia Fernando 
Published: June 2008

ABSTRACT

Many pathogenic bacteria subvert normal host cell processes by delivering effector proteins which mimic eukaryotic functions directly into target cells. EspF is a multifunctional protein injected into host cells by attaching and effacing pathogens, but its mechanism of action is not understood completely. In silico analyses of EspF revealed two key motifs: proline-rich domains and PDZ domain binding motifs. Such functional domains may allow EspF to act as an actin nucleation-promoting factor by mimicking host proteins. In agreement with these predictions, we found that EspF from rabbit enteropathogenic
Escherichia coli
(E22) participates in the regulation of actin polymerization by binding to a complex of proteins at the tight junctions (TJ). EspF bound to actin and profilin throughout the course of infection. However, after 2 h of infection, EspF also bound to the neural Wiskott-Aldrich syndrome protein and to the Arp2/3, zonula occludens-1 (ZO-1), and ZO-2 proteins. Moreover, EspF caused occludin, claudin, ZO-1, and ZO-2 redistribution and loss of transepithelial electrical resistance, suggesting that actin sequestration by EspF may cause local actin depolymerization leading to EspF-induced TJ disruption. Furthermore, EspF caused recruitment of these TJ proteins into the pedestals. An E22 strain lacking EspF did not cause TJ disruption and pedestals were smaller than those induced by the wild-type strain. Additionally, the pedestals were located mainly in the TJ. The overexpression of EspF caused bigger pedestals located along the length of the cells. Thus, actin sequestration by EspF allows the recruitment of junctional proteins into the pedestals, leading to the maturation of actin pedestals and the disruption of paracellular permeability.
A
Listeria monocytogenes
Mutant Defective in Bacteriophage Attachment Is Attenuated in Orally Inoculated Mice and Impaired in Enterocyte Intracellular Growth
Authors: Spears Patricia A. , Suyemoto M. Mitsu , Palermo Angela M. , Horton John R. , Hamrick Terri S. , Havell Edward A. , Orndorff Paul E. 
Published: June 2008

ABSTRACT

A
Listeria monocytogenes
bacteriophage was used to identify a phage-resistant Tn
917
insertion mutant of the mouse-virulent listerial strain F6214-1. The mutant was attenuated when it was inoculated orally into female A/J mice and failed to replicate efficiently in cultured mouse enterocytes. Phage binding studies indicated that the mutant had a cell surface alteration that precluded phage attachment. All phenotypes associated with the mutation could be complemented in
trans
by a single open reading frame (ORF) that corresponded to the ORF interrupted by the Tn
917
insertion. The complementation effected was, in all cases, at a level indistinguishable from that of the parent. The Tn
917
insertion interrupted a gene that is predicted to encode a group 2 glycosyl transferase (provisionally designated
glcV
). A similar
glcV
gene is present in
Listeria welshimeri
and
Listeria innocua
and in some serotypes of
L. monocytogenes
. We speculate that the loss of the
glcV
product results in a defective phage receptor and that this alteration coincidentally influences a feature of the normal host-pathogen interaction required for virulence. Interestingly, the
glcV
lesion, while preventing phage attachment, did not alter the mutant's ability to bind to cultured mouse enterocyte monolayers. Rather, the mutation appeared to alter a subsequent step in intracellular replication measured by a reduction in plaque-forming efficiency and plaque size. In vivo, the mutant was undetectable in the liver and spleen 48 h after oral inoculation. The mutation is significant in part because it is one of the few that produce attenuation when the mutant is delivered orally.
Alterations of Splenic Architecture in Malaria Are Induced Independently of Toll-Like Receptors 2, 4, and 9 or MyD88 and May Affect Antibody Affinity
Authors: Cadman Emma T. , Abdallah Asmahan Y. , Voisine Cécile , Sponaas Anne-Marit , Corran Patrick , Lamb Tracey , Brown Douglas , Ndungu Francis , Langhorne Jean 
Published: June 2008

ABSTRACTSplenic microarchitecture is substantially altered during acute malaria infections, which may affect the development and regulation of immune responses. Here we investigated whether engagement of host Toll-like receptor 2 (TLR2), TLR4, TLR9, and the adaptor protein MyD88 is required for induction of the changes and whether antibody responses are modified when immunization takes place during the period of splenic disruption. The alterations in splenic microarchitecture were maximal shortly after the peak of parasitemia and were not dependent on engagement of TLR2, TLR4, or TLR9, and they were only minimally affected by the absence of the MyD88 adaptor molecule. Although germinal centers were formed in infected mice, they did not contain the usual light and dark zones. Immunization of mice with chicken gamma globulin 2 weeks prior to acutePlasmodium chabaudiinfection did not affect the quantity or avidity of the immunoglobulin G antibody response to this antigen. However, immunization at the same time as the primaryP. chabaudiinfection resulted in a clear transient reduction in antibody avidity in the month following immunization. These data suggest that the alterations in splenic structure, particularly the germinal centers, may affect the quality of an antibody response during a malaria infection and could impact the development of immunity to malaria or to other infections or immunizations given during a malaria infection.