bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intrinsic Resistance of
Mycobacterium
smegmatis
to Fluoroquinolones May Be Influenced by New Pentapeptide Protein MfpA
Authors: Montero Clemente , Mateu Guaniri , Rodriguez Rosalva , Takiff Howard 
Published: July 2002

ABSTRACT

The fluoroquinolones (FQ) are used in the treatment of
Mycobacterium
tuberculosis
, but the development of resistance could limit their effectiveness. FQ resistance (FQ
R
) is a multistep process involving alterations in the type II topoisomerases and perhaps in the regulation of efflux pumps, but several of the steps remain unidentified. Recombinant plasmid pGADIV was selected from a genomic library of wild-type (WT), FQ-sensitive
M.
smegmatis
by its ability to confer low-level resistance to sparfloxacin (SPX). In WT
M
.
smegmatis
, pGADIV increased the MICs of ciprofloxacin (CIP) by fourfold and of SPX by eightfold, and in
M
.
bovis
BCG it increased the MICs of both CIP and SPX by fourfold. It had no effect on the accumulation of
14
C-labeled CIP or SPX. The open reading frame responsible for the increase in FQ
R
,
mfpA
, encodes a putative protein belonging to the family of pentapeptides, in which almost every fifth amino acid is either leucine or phenylalanine. Very similar proteins are also present in
M
.
tuberculosis
and
M
.
avium
. The MICs of CIP and SPX were lower for an
M
.
smegmatis
mutant strain lacking an intact
mfpA
gene than for the WT strain, suggesting that, by some unknown mechanism, the gene product plays a role in determining the innate level of FQ
R
in
M
.
smegmatis
.
Structural and Functional Analysis of Horse Cathelicidin Peptides
Authors: Skerlavaj Barbara , Scocchi Marco , Gennaro Renato , Risso Angela , Zanetti Margherita 
Published: July 2002

ABSTRACTCathelicidin-derived antimicrobial peptides are a component of the peptide-based host defense of neutrophils and epithelia, with a widespread distribution in mammals. We recently reported the cDNA sequences of three putative horse myeloid cathelicidins, named eCATH-1, -2, and -3. A Western analysis was performed to investigate their presence in neutrophils and processing to mature peptides. eCATH-2 and eCATH-3, but not eCATH-1, were found to be present in uncleaved forms in horse neutrophils. The corresponding mature peptides were detected in inflammatory sites, suggesting that processing of the propeptides takes place upon neutrophil activation. A functional characterization was then performed with synthetic eCATH peptides. Circular dichroism measurements indicated an amphipathic α-helical conformation of these peptides in an anisotropic environment, and in vitro assays revealed a potent activity and a broad spectrum of antimicrobial activity for eCATH-1 and a somewhat more restricted spectrum of activity for eCATH-2. Conversely, a strong dependence on salt concentration was observed when the activity of eCATH-3 was tested. This peptide efficiently killed bacteria and some fungal species, i.e.,Cryptococcus neoformansandRhodotorula rubra, in low-ionic-strength media, but the activity was inhibited in the presence of physiological salt medium. This behavior could be modified by modulating the amphipathicity of the molecule. In fact, the synthetic analogue LLK-eCATH-3, with a slightly modified sequence that increases the hydrophobic moment of the peptide, displayed a potent activity in physiological salt medium against the strains resistant to eCATH-3 under these conditions.
Pharmacokinetics of Ethionamide Administered under Fasting Conditions or with Orange Juice, Food, or Antacids
Authors: Auclair Barbara , Nix David E. , Adam Rodney D. , James Gordon T. , Peloquin Charles A. 
Published: July 2002

ABSTRACTThis study was conducted in order to (i) determine the effect of food, orange juice, or antacids on the absorption of a single oral 500-mg dose of ethionamide (ETA) in healthy volunteers, including an assessment of bioequivalence, and (ii) determine ETA population pharmacokinetic (PK) parameters. The pharmacokinetics of ETA in serum was determined for 12 healthy males and females in a randomized, four-period crossover study. Volunteers received single 500-mg doses of ETA either on an empty stomach (reference) or with food, orange juice, or antacids. Serum samples were collected for 48 h and assayed by high-performance liquid chromatography. Data were analyzed by noncompartmental and population methods. Mean test/reference ratios and 90% confidence intervals were determined. No statistically significant differences were seen in the maximum concentration of ETA (Cmax), time to maximum concentration (Tmax), or area under the concentration-time curve from 0 h to infinity (AUC0–∞) between the four treatments (P> 0.05 by analysis of variance). The least-squares mean ratios (with confidence intervals in parentheses) forCmaxwere 105% (81.2 to 135%) after orange juice, 94% (72.8 to 121%) after food, and 88% (68.4 to 114%) after antacids. The least-squares mean ratios (with confidence intervals is in parentheses) for AUC0–∞were 91% (72.7 to 115%) after orange juice, 96% (76.4 to 121%) after food, and 95% (75.5 to 120%) after antacids. The meanTmaxwas slightly prolonged following antacid or food administration (2.3 to 2.6 h) compared to administration on an empty stomach or with juice (1.7 to 1.9 h). The median population PK parameters were as follows:Ka= 0.37 to 0.48 h−1,V/F= 2.0 to 2.8 liters/kg, CL/F= 56.5 to 72.2 liters/h, and terminal half-life = 1.7 to 2.1 h, whereKais the absorption rate constant,Vis the volume of distribution, and CL is clearance. The PK behavior of ETA was not significantly modified by the different conditions studied. Mean ratios for AUC ranged from 0.91 to 0.96 for the orange juice, food, and antacid treatments, indicating a minimal effect on relative bioavailability. ETA can, therefore, be administered with food if tolerance is an issue.
Variant Human Immunodeficiency Virus Type 1 Proteases and Response to Combination Therapy Including a Protease Inhibitor
Authors: Servais Jean , Lambert Christine , Fontaine Elodie , Plesséria Jean-Marc , Robert Isabelle , Arendt Vic , Staub Thérèse , Schneider François , Hemmer Robert , Burtonboy Guy , Schmit Jean-Claude 
Published: July 2002

ABSTRACTThe objective of this observational study was to assess the genetic variability in the human immunodeficiency virus (HIV) protease gene from HIV type 1 (HIV-1)-positive (clade B), protease inhibitor-naı̈ve patients and to evaluate its association with the subsequent effectiveness of a protease inhibitor-containing triple-drug regimen. The protease gene was sequenced from plasma-derived virus from 116 protease inhibitor-naı̈ve patients. The virological response to a triple-drug regimen containing indinavir, ritonavir, or saquinavir was evaluated every 3 months for as long as 2 years (n= 40). A total of 36 different amino acid substitutions compared to the reference sequence (HIV-1 HXB2) were detected. No substitutions at the active site similar to the primary resistance mutations were found. The most frequent substitutions (prevalence, >10%) at baseline were located at codons 15, 13, 12, 62, 36, 64, 41, 35, 3, 93, 77, 63, and 37 (in ascending order of frequency). The mean number of polymorphisms was 4.2. A relatively poorer response to therapy was associated with a high number of baseline polymorphisms and, to a lesser extent, with the presence of I93L at baseline in comparison with the wild-type virus. A71V/T was slightly associated with a poorer response to first-line ritonavir-based therapy. In summary, within clade B viruses, protease gene natural polymorphisms are common. There is evidence suggesting that treatment response is associated with this genetic background, but most of the specific contributors could not be firmly identified. I93L, occurring in about 30% of untreated patients, may play a role, as A71V/T possibly does in ritonavir-treated patients.
Penicillin-Binding Protein 5 and Expression of Ampicillin Resistance in
Enterococcus faecium
Authors: Rice Louis B. , Carias Lenore L. , Hutton-Thomas Rebecca , Sifaoui Farid , Gutmann Laurent , Rudin Susan D. 
Published: July 2002

ABSTRACT

We report a structural and transcriptional analysis of the
pbp5
region of
Enterococcus faecium
C68.
pbp5
exists within a larger operon that includes upstream open reading frames (ORFs) corresponding to previously reported
psr
(penicillin-binding protein synthesis repressor) and
ftsW
(whose product is a transmembrane protein that interacts with PBP3 in
Escherichia coli
septum formation) genes. Hybridization of mRNA from C68, CV133, and four ampicillin-resistant CV133 mutants revealed four distinct transcripts from this region, consisting of (i)
E. faecium ftsW
(

ftsW
Efm

) alone; (ii)
psr
and
pbp5
; (iii)
pbp5
alone; and (iv)

ftsW
Efm

,
psr
, and
pbp5
. Quantities of the different transcripts varied between strains and did not always correlate with quantities of PBP5 or levels of ampicillin resistance. Since the
psr
of C68 is presumably nonfunctional due to an insertion of an extra nucleotide in the codon for the 44th amino acid, the region extending from the

ftsW
Efm

promoter through the
pbp5
gene of C68 was cloned in
E. coli
to facilitate mutagenesis. The
psr
ORF was regenerated using site-directed mutagenesis and introduced into
E. faecium
D344-SRF on conjugative shuttle vector pTCV-lac (pCWR558 [
psr
ORF interrupted]; pCWR583 [
psr
ORF intact]). Ampicillin MICs for both D344-SRF(pCWR558) and D344-SRF(pCWR583) were 64 μg/ml. Quantities of
pbp5
transcript and protein were similar in strains containing either construct regardless of whether they were grown in the presence or absence of ampicillin, arguing against a role for PSR as a repressor of
pbp5
transcription. However, quantities of
psr
transcript were increased in D344-SRF(pCWR583) compared to D344-SRF(pCWR558), especially after growth in ampicillin; suggesting that PSR acts in some manner to activate its own transcription.
Cross-Resistance Testing of Antihepadnaviral Compounds Using Novel Recombinant Baculoviruses Which Encode Drug-Resistant Strains of Hepatitis B Virus
Authors: Delaney William E. , Edwards Ros , Colledge Danni , Shaw Tim , Torresi Joseph , Miller Thomas G. , Isom Harriet C. , Bock C. Thomas , Manns Michael P. , Trautwein Christian , Locarnini Stephen 
Published: July 2002

ABSTRACTLong-term nucleoside analog therapy for hepatitis B virus (HBV)-related disease frequently results in the selection of mutant HBV strains that are resistant to therapy. Molecular studies of such drug-resistant variants are clearly warranted but have been difficult to do because of the lack of convenient and reliable in vitro culture systems for HBV. We previously developed a novel in vitro system for studying HBV replication that relies on the use of recombinant baculoviruses to deliver greater than unit length copies of the HBV genome to HepG2 cells. High levels of HBV replication can be achieved in this system, which has recently been used to assess the effects of lamivudine on HBV replication and covalently closed circular DNA accumulation. The further development of this novel system and its application to determine the cross-resistance profiles of drug-resistant HBV strains are described here. For these studies, novel recombinant HBV baculoviruses which encoded the L526M, M550I, and L526M M550V drug resistance mutations were generated and used to examine the effects of these substitutions on viral sensitivity to lamivudine, penciclovir (the active form of famciclovir), and adefovir, three compounds of clinical importance. The following observations were made: (i) the L526M mutation confers resistance to penciclovir and partial resistance to lamivudine, (ii) the YMDD mutations M550I and L526M M550V confer high levels of resistance to lamivudine and penciclovir, and (iii) adefovir is active against each of these mutants. These findings are supported by the limited amount of clinical data currently available and confirm the utility of the HBV-baculovirus system as an in vitro tool for the molecular characterization of clinically significant HBV strains.
In Vitro Activities of Linezolid, Meropenem, and Quinupristin-Dalfopristin against Group C and G Streptococci, Including Vancomycin-Tolerant Isolates
Authors: Zaoutis Theoklis , Moore Lynn Steele , Furness Kathleen , Klein Joel D. 
Published: July 2002

ABSTRACT
The in vitro activities of meropenem, linezolid, quinupristin-dalfopristin, vancomycin, and penicillin against 130 clinical isolates of group C and G streptococci, including vancomycin-tolerant isolates, were evaluated. Meropenem, linezolid, quinupristin-dalfopristin, vancomycin, and penicillin MICs at which 90% of the isolates were inhibited were 0.06, 2.0, 0.25, 0.5, and ≤0.016 μg/ml, respectively. Meropenem, linezolid, quinupristin-dalfopristin, and penicillin were active against group C and G streptococci, including vancomycin-resistant strains.
Efficacy of Linezolid in Treatment of Experimental Endocarditis Caused by Methicillin-Resistant
Staphylococcus aureus
Authors: Dailey Charlene F. , Dileto-Fang Christine L. , Buchanan Lewis V. , Oramas-Shirey Martha P. , Batts Donald H. , Ford Charles W. , Gibson John K. 
Published: July 2002

ABSTRACT

The efficacies of orally (p.o.) dosed linezolid and intravenously (i.v.) dosed vancomycin against methicillin-resistant
Staphylococcus aureus
(MRSA) in rabbits with experimental aortic-valve endocarditis were investigated. After endocarditis was established with a recent clinical MRSA isolate, rabbits were dosed for 5 days with linezolid (p.o., three times a day) at either 25, 50, or 75 mg/kg of body weight or vancomycin (i.v., twice a day) at 25 mg/kg. The 25-mg/kg linezolid group had a high mortality rate and bacterial counts in the valve vegetations that were not different from those of the controls. Linezolid dosed p.o. at 50 and 75 mg/kg and i.v. vancomycin produced statistically significant reductions in bacterial counts compared to those of the untreated controls. The reduced bacterial counts and culture-negative valve rates for the animals treated with linezolid at 75 mg/kg were similar to those for the vancomycin-treated animals. Concentrations of linezolid in plasma were determined at several points in the dosing regimen. These results suggest that the efficacy of linezolid in this infection model is related to trough levels in plasma that remain above the MIC for this microorganism. At the ineffective dose of linezolid (25 mg/kg) the concentration at sacrifice was 0.045 times the MIC, whereas the concentrations of linezolid in plasma in the 50- and 75-mg/kg groups were 2 and 5 times the MIC at sacrifice, respectively. The results from this experimental model suggest that the oxazolidinone linezolid may be effective for the treatment of serious staphylococcal infections when resistance to other antimicrobials is present.
Mycobacterium smegmatis
d
-Alanine Racemase Mutants Are Not Dependent on
d
-Alanine for Growth
Authors: Chacon Ofelia , Feng Zhengyu , Harris N. Beth , Cáceres Nancy E. , Adams L. Garry , Barletta Raúl G. 
Published: July 2002

ABSTRACT

Mycobacterium smegmatis
is a fast-growing nonpathogenic species particularly useful in studying basic cellular processes of relevance to pathogenic mycobacteria. This study focused on the
d
-alanine racemase gene (
alrA
), which is involved in the synthesis of
d
-alanine, a basic component of peptidoglycan that forms the backbone of the cell wall.
M. smegmatis alrA
null mutants were generated by homologous recombination using a kanamycin resistance marker for insertional inactivation. Mutants were selected on Middlebrook medium supplemented with 50 mM
d
-alanine and 20 μg of kanamycin per ml. These mutants were also able to grow in standard and minimal media without
d
-alanine, giving rise to colonies with a drier appearance and more-raised borders than the wild-type strain. The viability of the mutants and independence of
d
-alanine for growth indicate that inactivation of
alrA
does not impose an auxotrophic requirement for
d
-alanine, suggesting the existence of a new pathway of
d
-alanine biosynthesis in
M. smegmatis
. Biochemical analysis demonstrated the absence of any detectable
d
-alanine racemase activity in the mutant strains. In addition, the
alrA
mutants displayed hypersusceptibility to the antimycobacterial agent
d
-cycloserine. The MIC of
d
-cycloserine for the mutant strain was 2.56 μg/ml, 30-fold less than that for the wild-type strain. Furthermore, this hypersusceptibility was confirmed by the bactericidal action of
d
-cycloserine on broth cultures. The kinetic of killing for the mutant strain followed the same pattern as that for the wild-type strain, but at a 30-fold-lower drug concentration. This effect does not involve a change in the permeability of the cell wall by this drug and is consistent with the identification of
d
-alanine racemase as a target of
d
-cycloserine. This outcome is of importance for the design of novel antituberculosis drugs targeting peptidoglycan biosynthesis in mycobacteria.
Penetration of Rifampin through
Staphylococcus epidermidis
Biofilms
Authors: Zheng Zhilan , Stewart Philip S. 
Published: July 2002

ABSTRACT

Rifampin penetrated biofilms formed by
Staphylococcus epidermidis
but failed to effectively kill the bacteria. Penetration was demonstrated by a simple diffusion cell bioassay and by transmission electron microscopic observation of antibiotic-affected cells at the distal edge of the biofilm.
In Vitro Interactions of Artemisinin with Atovaquone, Quinine, and Mefloquine againstPlasmodium falciparum
Authors: Gupta S. , Thapar M. M. , Wernsdorfer W. H. , Björkman A. 
Published: July 2002

ABSTRACTThe interactions of artemisinin with atovaquone, quinine, and mefloquine were investigated in threePlasmodium falciparumstrains (strains F-32, FCR-3, and K-1) by an in vitro culture assay. The parasites were cultured for 48 h in the presence of different concentrations and proportions of two drugs at a time in a checkerboard design. The response parameters were determined, and the sums of the fractional inhibitory concentrations (ΣFICs) of the drug combinations were calculated for different degrees of inhibition (50% effective concentration [EC50], EC90, and EC99). Within therapeutically relevant molar ratios (19 to 200), the combination of quinine and artemisinin showed mean ΣFICs of 1.71 at the EC50, 0.36 at the EC90, and 0.13 at the EC99, indicating increasing synergism. Within the range of molar ratios of 4.3 to 50, the combination of mefloquine and artemisinin yielded mean ΣFCIs of 0.93, 0.44, and 0.31 at the EC50, EC90, and EC99, respectively, indicating synergism. The atovaquone combination showed additive activity to synergism at atovaquone/artemisinin proportions considered relevant to the in vivo situation, i.e., between 4.3 and 200, with the mean ΣFICs decreasing from 1.34 at the EC50to 0.85 and 0.23 at the EC90and EC99, respectively. Interstrain differences in the degree of drug interaction were seen with the three strains for all combinations. Synergism was most consistent with quinine.
Shiga-Like Toxin II Derived fromEscherichia coliO157:H7 Modifies Renal Handling of Levofloxacin in Rats
Authors: Zhao Ying Lan , Cen Xiao Bo , Ito Masafumi , Yokoyama Keiko , Takagi Kenji , Kitaichi Kiyoyuki , Nadai Masayuki , Ohta Michio , Takagi Kenzo , Hasegawa Takaaki 
Published: July 2002

ABSTRACTThe effect of Shiga-like toxin II (SLT-II) (2 μg/animal), which was derived fromEscherichia coliO157:H7, on renal handling of levofloxacin (LVX), a model drug for quinolone antimicrobial agents, was investigated in rats 24 h after intravenous injection. In histopathological examination, acute tubular injury was observed in SLT-II-treated rats, but the glomeruli were not injured. SLT-II significantly increased the steady-state concentration of LVX in plasma to 1.5-fold that of control rats. SLT-II induced significant decreases in the glomerular filtration rate (GFR) and renal clearance (CLR) of LVX. SLT-II slightly, but significantly, increased the unbound fraction and decreased renal plasma flow with no change in the extraction ratio ofp-aminohippurate. SLT-II significantly increased concentrations of tumor necrosis factor alpha (TNF-α) and nitrite and nitrate (NOx) in plasma. The TNF-α inhibitor pentoxifylline partly, but significantly, inhibited SLT-II-induced decreases in the GFR and CLRof LVX; in contrast,S-methylisothiourea, a selective inhibitor of inducible nitric oxide synthase, did not. Western blotting analysis revealed that SLT-II did not alter the levels of multidrug resistance-associated protein 2 (Mrp2) and P-glycoprotein in kidneys 24 h after injection, assuming the lack of involvement of Mrp2 and P-glycoprotein in SLT-II-induced acute renal tubular injury and renal handling of LVX observed 24 h after SLT-II injection. The present study suggests that SLT-II impairs the renal handling of LVX by decreasing GFR and causing decreased renal plasma flow.
Mechanism of Action ofMelaleuca alternifolia(Tea Tree) Oil onStaphylococcus aureusDetermined by Time-Kill, Lysis, Leakage, and Salt Tolerance Assays and Electron Microscopy
Authors: Carson Christine F. , Mee Brian J. , Riley Thomas V. 
Published: July 2002

ABSTRACTThe essential oil ofMelaleuca alternifolia(tea tree) has broad-spectrum antimicrobial activity. The mechanisms of action of tea tree oil and three of its components, 1,8-cineole, terpinen-4-ol, and α-terpineol, againstStaphylococcus aureusATCC 9144 were investigated. Treatment with these agents at their MICs and two times their MICs, particularly treatment with terpinen-4-ol and α-terpineol, reduced the viability ofS. aureus. None of the agents caused lysis, as determined by measurement of the optical density at 620 nm, although cells became disproportionately sensitive to subsequent autolysis. Loss of 260-nm-absorbing material occurred after treatment with concentrations equivalent to the MIC, particularly after treatment with 1,8-cineole and α-terpineol.S. aureusorganisms treated with tea tree oil or its components at the MIC or two times the MIC showed a significant loss of tolerance to NaCl. When the agents were tested at one-half the MIC, only 1,8-cineole significantly reduced the tolerance ofS. aureusto NaCl. Electron microscopy of terpinen-4-ol-treated cells showed the formation of mesosomes and the loss of cytoplasmic contents. The predisposition to lysis, the loss of 260-nm-absorbing material, the loss of tolerance to NaCl, and the altered morphology seen by electron microscopy all suggest that tea tree oil and its components compromise the cytoplasmic membrane.