bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Bacteriophage Lambda-Based Genetic Screen for Characterization of the Activity and Phenotype of the Human Immunodeficiency Virus Type 1 Protease
Authors: Martínez Miguel-Angel , Cabana Marta , Parera Mariona , Gutierrez Arantxa , Esté José A. , Clotet Bonaventura 
Published: July 2002

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) resistance to antiretroviral drugs is the main cause of patient treatment failure. Despite the problems associated with interpretation of HIV-1 resistance testing, resistance monitoring should help in the rational design of initial or rescue antiretroviral therapies. It has previously been shown that the activity of the HIV-1 protease can be monitored by using a bacteriophage lambda-based genetic assay. This genetic screening system is based on the bacteriophage lambda regulatory circuit in which the viral repressor
c
I is specifically cleaved to initiate the lysogenic to lytic switch. We have adapted this simple lambda-based genetic assay for the analysis of the activities and phenotypes of different HIV-1 proteases. Lambda phages that encode HIV-1 proteases either from laboratory strains (strain HXB2) or from clinical samples are inhibited in a dose-dependent manner by the HIV-1 protease inhibitors indinavir, ritonavir, saquinavir, and nelfinavir. Distinct susceptibilities to different drugs were also detected among phages that encode HIV-1 proteases carrying different resistance mutations, further demonstrating the specificity of this assay. Differences in proteolytic processing activity can also be directly monitored with this genetic screen system since two phage populations compete in culture with each other until one phage outgrows the other. In summary, we present here a simple, safe, and rapid genetic screening system that may be used to predict the activities and phenotypes of HIV-1 proteases in the course of viral infection and antiretroviral therapy. This assay responds appropriately to well-known HIV-1 protease inhibitors and can be used to search for new protease inhibitors.
Evaluation of the Invader Assay, a Linear Signal Amplification Method, for Identification of Mutations Associated with Resistance to Rifampin and Isoniazid in
Mycobacterium tuberculosis
Authors: Cooksey Robert C. , Holloway Brian P. , Oldenburg Mary C. , Listenbee Sonja , Miller Carolyn W. 
Published: July 2002

ABSTRACT

We evaluated a recently described linear signal amplification method for sensitivity and specificity in detecting mutations associated with resistance to rifampin (RIF) and isoniazid (INH) in
Mycobacterium tuberculosis
. The assay utilizes the thermostable flap endonuclease Cleavase VIII, derived from
Archaeoglobus fulgidus
, which cleaves a structure formed by the hybridization of two overlapping oligonucleotide probes to a target nucleic acid strand. This method, termed the Invader assay, can discriminate single-base differences. Nine pairs of probes, encompassing five mutations in
rpoB
and
katG
that are associated with resistance to either RIF or INH, as well as the corresponding wild-type (drug-susceptible) alleles, were tested using amplified DNA. Fluorescent-labeled cleavage products, ranging from 4 to 13 nucleotides in length, depending on the genotype of the test sample, were separated by denaturing polyacrylamide (20 to 24%) gel electrophoresis and then detected by scanning. All nine alleles could be identified and differentiated on the basis of product size. Multiple mutations at a specific
rpoB
nucleotide in target PCR products could be identified, as could mutants that were present at ≥0.5% of the total population of target sequences. The Invader assay is a sensitive screen for some mutations associated with antituberculosis drug resistance in amplified gene regions.
Comparative In Vitro Activities of Clinafloxacin and Trovafloxacin against 1,000 Isolates of
Bacteroides fragilis
Group: Effect of the Medium on Test Results
Authors: Snydman David R. , Jacobus Nilda V. , McDermott Laura A. , Supran Stacey E. 
Published: July 2002

ABSTRACT

The in vitro antibacterial activities of clinafloxacin, trovafloxacin, ciprofloxacin, and cefoxitin against 1,000 clinical isolates of
Bacteroides fragilis
group were compared by agar dilution in brucella blood agar (BBA) and Wilkins Chalgren agar (WCA). Significantly higher geometric mean MICs for the three quinolones and cefoxitin (
P
< 0.001) were obtained in BBA than in WCA. Regardless of medium, clinafloxacin was slightly more active than trovafloxacin. The activity of clinafloxacin and trovafloxacin was greater than that of cefoxitin against
B. distasonis
,
B. ovatus
, and
B. thetaiotaomicron
but lower against
B. vulgatus
. High cross resistance between trovafloxacin and clinafloxacin was observed.
In Vitro Activities of Voriconazole, Itraconazole, and Amphotericin B against
Blastomyces dermatitidis
,
Coccidioides immitis
, and
Histoplasma capsulatum
Authors: Li Ren-Kai , Ciblak Meral A. , Nordoff Nicole , Pasarell Lester , Warnock David W. , McGinnis Michael R. 
Published: July 2002

ABSTRACT

The in vitro activity of voriconazole was compared to those of itraconazole and amphotericin B against the mold forms of 304 isolates of three dimorphic fungi,
Blastomyces dermatitidis
,
Coccidioides immitis
, and
Histoplasma capsulatum
. MICs were determined by a broth microdilution adaptation of the National Committee for Clinical Laboratory Standards M27-A procedure. RPMI 1640 medium was used for tests with voriconazole and itraconazole, whereas Antibiotic Medium 3 with 2% glucose was used for amphotericin B. Minimum fungicidal concentrations (MFCs) were also determined. Amphotericin B was active against all three dimorphic fungi, with MICs at which 90% of the isolates tested are inhibited (MIC
90
s) of 0.5 to 1 μg/ml. Itraconazole had MIC
90
s of 0.06 μg/ml for
H. capsulatum
, 0.125 μg/ml for
B. dermatitidis
, and 1 μg/ml for
C. immitis
. The MIC
90
s of voriconazole were 0.25 μg/ml for all three fungi. Amphotericin B was fungicidal for
B. dermatitidis
and
H. capsulatum
with MFCs at which 90% of strains tested are killed (MFC
90
s) of 0.5 and 2 μg/ml, respectively. It was less active against
C. immitis
, with MFCs ranging from 0.5 to >16 μg/ml. Voriconazole and itraconazole were lethal for most isolates of
B. dermatitidis
, with MFC
50
s and MFC
90
s of 0.125 and 4 μg/ml, respectively. Both azoles were fungicidal for some isolates of
H. capsulatum
, with MFC
50
s of 2 and 8 μg/ml for itraconazole and voriconazole, respectively; neither had a lethal effect upon
C. immitis
. Our results suggest that voriconazole possesses promising activity against these important human pathogens.
Antimicrobial Susceptibilities and Clinical Sources of
Facklamia
Species
Authors: LaClaire L. , Facklam R. 
Published: July 2002

ABSTRACT

Facklamia
spp. are gram-positive cocci, arranged in short chains or diplos, and resemble viridans streptococci on 5% sheep blood agar. Eighteen strains representing four species of
Facklamia
were isolated from blood cultures, an abscess, bone, cerebrospinal fluid, gall bladder, vaginal swab, and one unknown source. Cultures were tested against 15 antimicrobial agents by using the broth microdilution MIC method. Reduced susceptibilities to the beta lactams, erythromycin, clindamycin, trimethoprim-sulfamethoxazole, and tetracycline were found. These results indicate that the susceptibilities of the
Facklamia
species are varied and that some strains have resistance patterns which may present difficulty in managing systemic infections in patients.
Contribution of a Thickened Cell Wall and Its Glutamine Nonamidated Component to the Vancomycin Resistance Expressed by
Staphylococcus aureus
Mu50
Authors: Cui Longzhu , Murakami Hiroko , Kuwahara-Arai Kyoko , Hanaki Hideaki , Hiramatsu Keiichi 
Published: July 2002

ABSTRACT

Staphylococcus aureus
Mu50, which has reduced susceptibility to vancomycin, has a remarkably thickened cell wall with an increased proportion of glutamine nonamidated muropeptides. In addition, Mu50 had enhanced glutamine synthetase and
l
-glutamine
d
-fructose-6-phosphate aminotransferase activities, which are involved in the cell-wall peptidoglycan synthesis pathway. Furthermore, significantly increased levels of incorporation of
14
C-labeled
d
-glucose into the cell wall was observed in Mu50. Unlike a
femC
mutant
S. aureus
strain, increased levels of production of nonamidated muropeptides in Mu50 was not caused by lower levels of glutamine synthetase activity but was considered to be due to the glutamine depletion caused by increased glucose utilization by the cell to biosynthesize increased amounts of peptidoglycan. After the cells were allowed to synthesize cell wall in the absence or presence of glucose and glutamine, cells with different cell-wall thicknesses and with cell walls with different levels of cross-linking were prepared, and susceptibility testing of these cells demonstrated a strong correlation between the cell-wall thickness and the degree of vancomycin resistance. Affinity trapping of vancomycin molecules by the cell wall and clogging of the outer layers of peptidoglycan by bound vancomycin molecules were considered to be the mechanism of vancomycin resistance of Mu50. The reduced cross-linking and the increased affinity of binding to vancomycin of the Mu50 cell wall presumably caused by the increased proportion of nonamidated muropeptides may also contribute to the resistance to some extent.
Accumulation of Norfloxacin by
Bacteroides fragilis
Authors: Ricci Vito , Piddock Laura J. V. 
Published: July 2002

ABSTRACT

The accumulation of norfloxacin by
Bacteroides fragilis
NCTC 9343 was determined by the modified fluorescence method. The time required to achieve a steady-state concentration (SSC) after allowing
B. fragilis
to accumulate norfloxacin in an aerobic or an anaerobic environment was ∼2 min; the SSC achieved in air was 90.28 ± 9.32 ng of norfloxacin/mg (dry weight) of cells, and that achieved anaerobically was 98.45 ± 3.7 ng of norfloxacin/mg (dry weight) of cells. Initial rates of accumulation were determined with a range of external concentrations, as up to 8 μg/ml the concentration of norfloxacin accumulated increased proportionally to the external concentration, 12.13 ng/mg (dry weight) of cells per μg of exogenous norfloxacin per ml. At concentrations above 10 μg/ml no increase in the rate of norfloxacin accumulation was observed. From the kinetic data, a Lineweaver-Burk plot calculated a

K
m

of 5.03 μg/ml and a
V
max
of 25.1 ng of norfloxacin/s. With an increase in temperature of between 0 and 30°C, the concentration of norfloxacin accumulated also increased proportionally at 4.722 ng of norfloxacin/mg (dry weight) of cells/°C. At low concentrations of glucose (<0.2%; 11 mM), the concentration of norfloxacin accumulated was decreased. With the addition of 100 μM carbonyl cyanide
m
-chlorophenylhydrazone (CCCP) the mean SSC of norfloxacin was increased to 116 ± 7.01 ng of norfloxacin/mg (dry weight) of cells; glucose had no significant effect in the presence of CCCP. Magnesium chloride (20 mM) decreased the SSC of norfloxacin to 40.5 ± 3.76 ng of norfloxacin per mg (dry weight) of cells. These data suggest that the mechanism of accumulation of norfloxacin by
B. fragilis
is similar to that of aerobic bacteria and that the fluoresence procedure is suitable for use with an anaerobic bacterium.
Trovafloxacin Treatment of Viridans Group Streptococcus Experimental Endocarditis
Authors: Piper Kerryl E. , Rouse Mark S. , Ronningen Karen L. , Steckelberg James M. , Wilson Walter R. , Patel Robin 
Published: July 2002

ABSTRACT

The activity of trovafloxacin was compared with those of vancomycin and penicillin in a model of
Streptococcus sanguis
species group (trovafloxacin MIC, 0.125 μg/ml) and
Streptococcus mitis
species group (trovafloxacin MIC, 0.125 μg/ml) experimental endocarditis. Rabbits with catheter-induced aortic valve vegetations were given no treatment, trovafloxacin at 15 mg/kg of body weight three times a day (t.i.d.), vancomycin at 15 mg/kg twice a day, or penicillin at 1.2 × 10
6
IU t.i.d. After 3 days of treatment, the animals were sacrificed; cardiac valve vegetations were aseptically removed and cultured quantitatively. Penicillin was as active as vancomycin as measured by in vivo clearance of bacteria. Trovafloxacin was less active (
P
< 0.05) than vancomycin or penicillin against
S. sanguis
species group infection but had similar efficacy against
S. mitis
species group infection. Quinolones, despite MICs in the susceptible range, may not be active for serious infections caused by some viridans group streptococci.
Validation of a Noninvasive, Real-Time Imaging Technology Using Bioluminescent
Escherichia coli
in the Neutropenic Mouse Thigh Model of Infection
Authors: Rocchetta H. L. , Boylan C. J. , Foley J. W. , Iversen P. W. , LeTourneau D. L. , McMillian C. L. , Contag P. R. , Jenkins D. E. , Parr T. R. 
Published: July 2002

ABSTRACT

A noninvasive, real-time detection technology was validated for qualitative and quantitative antimicrobial treatment applications. The
lux
gene cluster of
Photorhabdus luminescens
was introduced into an
Escherichia coli
clinical isolate, EC14, on a multicopy plasmid. This bioluminescent reporter bacterium was used to study antimicrobial effects in vitro and in vivo, using the neutropenic-mouse thigh model of infection. Bioluminescence was monitored and measured in vitro and in vivo with an intensified charge-coupled device (ICCD) camera system, and these results were compared to viable-cell determinations made using conventional plate counting methods. Statistical analysis demonstrated that in the presence or absence of antimicrobial agents (ceftazidime, tetracycline, or ciprofloxacin), a strong correlation existed between bioluminescence levels and viable cell counts in vitro and in vivo. Evaluation of antimicrobial agents in vivo could be reliably performed with either method, as each was a sound indicator of therapeutic success. Dose-dependent responses could also be detected in the neutropenic-mouse thigh model by using either bioluminescence or viable-cell counts as a marker. In addition, the ICCD technology was examined for the benefits of repeatedly monitoring the same animal during treatment studies. The ability to repeatedly measure the same animals reduced variability within the treatment experiments and allowed equal or greater confidence in determining treatment efficacy. This technology could reduce the number of animals used during such studies and has applications for the evaluation of test compounds during drug discovery.
Identification of an
erm
(A) Erythromycin Resistance Methylase Gene in
Streptococcus pneumoniae
Isolated in Greece
Authors: Syrogiannopoulos George A. , Grivea Ioanna N. , Tait-Kamradt Amelia , Katopodis George D. , Beratis Nicholas G. , Sutcliffe Joyce , Appelbaum Peter C. , Davies Todd A. 
Published: July 2002

ABSTRACT

In a serotype 11A clone of erythromycin-resistant pneumococci isolated from young Greek carriers, we identified the nucleotide sequence of
erm
(A), a methylase gene previously described as
erm
(TR) in
Streptococcus pyogenes
. The
erm
(A) pneumococci were resistant to 14- and 15-member macrolides, inducibly resistant to clindamycin, and susceptible to streptogramin B. To our knowledge, this is the first identification of resistance to erythromycin in
S. pneumoniae
attributed solely to the carriage of the
erm
(A) gene.
Cellular Uptake of Two Fluoroketolides, HMR 3562 and HMR 3787, by Human Polymorphonuclear Neutrophils In Vitro
Authors: Abdelghaffar H. , Vazifeh D. , Labro M. T. 
Published: July 2002

ABSTRACT

We analyzed the cellular accumulation of two new fluoroketolides, HMR 3562 and HMR 3787, by human polymorphonuclear neutrophils (PMN) in vitro. Both compounds were rapidly taken up by PMN, with a cellular-to-extracellular concentration ratio (C/E) of about 141 (HMR 3562) and 117 (HMR 3787) at 5 min, and this was followed by a plateau at 60 to 180 min, with a C/E of >300 at 180 min. Both ketolides were mainly located in PMN granules (about 75%) and egressed slowly from loaded cells (about 40% at 60 min), owing to avid reuptake. Uptake was moderately sensitive to external pH, and activation energy was also moderate (about 70 kJ/mol). As with other macrolides and ketolides, the existence of an active transport system was suggested by (i) the strong interindividual variability in uptake kinetics, suggesting variability in the number or activity of a transport protein; (ii) the saturation kinetics characteristic of a carrier-mediated transport system (
V
max
, about 2,300 ng/2.5 × 10
6
PMN/5 min;

K
m

, about 50 μg/ml); (iii) the inhibitory effects of Ni
2+
(a blocker of the Na
+
-Ca
2+
exchanger), phorbol myristate acetate (a protein kinase C activator), and H89 (a protein kinase A inhibitor). Although these two ketolides are more related to HMR 3647 (telithromycin), it is interesting that the presence of a fluoride gave these molecules a cellular pharmacokinetics more like those of HMR 3004 than those of HMR 3647. The macrolide transport system has not been yet elucidated, but our data confirm that, despite variations in chemical structure, all erythromycin A derivatives share a transmembrane transport system.
Transfer of Tn
916
-Like Elements in Microcosm Dental Plaques
Authors: Roberts Adam P. , Cheah Grace , Ready Derren , Pratten Jonathan , Wilson Michael , Mullany Peter 
Published: July 2002

ABSTRACT

Microcosm dental plaques were grown from an inoculum of human saliva in a constant-depth film fermentor. The inoculum contained four tetracycline-resistant streptococcal species, each of which contained a Tn
916
-like element. This element was shown to transfer to other streptococci both in filter-mating experiments and within the biofilms in the fermentor.
High Incidence of Erythromycin Resistance among Clinical Isolates of
Streptococcus agalactiae
in Taiwan
Authors: Hsueh Po-Ren , Teng Lee-Jene , Lee Li-Na , Ho Shen-Wu , Yang Pan-Chyr , Luh Kwen-Tay 
Published: July 2002

ABSTRACT

The in vitro susceptibilities of 266 isolates of
Streptococcus agalactiae
determined by the agar dilution method showed that 6% of isolates were nonsusceptible to penicillin and 46% was resistant to erythromycin. Of the erythromycin-resistant isolates, 86.3% had the macrolide-lincosamide-streptogramin (MLS) resistance phenotype (constitutive MLS, 85.5%; inducible MLS, 0.8%) and 13.7% had the M phenotype.
Fractional Maximal Effect Method for In Vitro Synergy between Amoxicillin and Ceftriaxone and between Vancomycin and Ceftriaxone against
Enterococcus faecalis
and Penicillin-Resistant
Streptococcus pneumoniae
Authors: Desbiolles Norbert , Piroth Lionel , Lequeu Catherine , Neuwirth Catherine , Portier Henri , Chavanet Pascal 
Published: July 2002

ABSTRACT

In the present study we assessed the use of a new in vitro testing method and graphical representation of the results to investigate the potential effectiveness of combinations of amoxicillin (AMZ) plus ceftriaxone (CRO) and of CRO plus vancomycin (VAN) against strains of
Streptococcus pneumoniae
highly resistant to penicillin and cephalosporins (PRP strains). We used the fractional maximal effect (FME) method of time-kill curves to calculate adequate concentrations of the drugs to be tested rather than relying on arbitrary choices. The concentrations obtained, each of which corresponded to a fraction of the maximal effect, were tested alone and in combination with the bacterial strains in a broth medium. Synergy was defined as a ratio of observed effect/theoretical effect, called FME, of greater than 1, additivity was defined as an FME equal to 1, and antagonism was defined as an observed effect lower than the best effect of one of the antibiotics used alone. The area between antagonism and additivity is the indifference zone. The well-known synergy between amoxicillin and gentamicin against a reference strain of
Enterococcus faecalis
was confirmed, with a best FME equal to 1.07. Two strains of PRP, strains PRP-1 and PRP-2, were studied. The MICs for PRP-1 and PRP-2 were as follows: penicillin, 4 and 16 μg/ml, respectively; AMZ, 2 and 8 μg/ml, respectively, CRO, 1 and 4 μg/ml, respectively; and VAN, 0.5 and 0.25 μg/ml, respectively. For PRP-1 the best FME for the combination AMZ-CRO was 1.22 with drug concentrations of 1.68 mg/liter for AMZ and 0.17 mg/liter for CRO; the best FME for the combination VAN-CRO was 1.75 with VAN at 0.57 mg/liter and CRO at 0.17 mg/liter. For PRP-2 the best FME obtained for the combination AMZ-CRO was 1.05 with drug concentrations of 11.28 mg/liter for AMZ and 0.64 mg/liter for CRO; the best FME obtained for the combination VAN-CRO was 1.35 with VAN at 0.25 mg/liter and CRO at 1.49 mg/liter. These results demonstrated the synergy of both combinations, AMZ-CRO and VAN-CRO, against PRP strains at drug concentrations achievable in humans. Consequently, either of the combinations can be proposed for use for the treatment of PRP infections.
Pharmacokinetic Interactions between Nelfinavir and 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Inhibitors Atorvastatin and Simvastatin
Authors: Hsyu Poe-Hirr , Schultz-Smith Melissa D. , Lillibridge James H. , Lewis Ronald H. , Kerr Bradley M. 
Published: July 2002

ABSTRACT

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are effective agents in lowering cholesterol and triglycerides and are being used by human immunodeficiency virus-positive patients to treat the lipid elevation that may be associated with antiretroviral therapy. Many HMG-CoA reductase inhibitors and protease inhibitors are metabolized by the same cytochrome P450 enzyme 3A4 (CYP3A4). In addition, many protease inhibitors are potent inhibitors of CYP3A4. Therefore, coadministration of these two classes of drugs may cause significant drug interactions. This open-label, multiple-dose study was performed to determine the interactions between nelfinavir, a protease inhibitor, and two HMG-CoA reductase inhibitors, atorvastatin and simvastatin, in healthy volunteers. Thirty-two healthy subjects received either atorvastatin calcium (10 mg once a day) or simvastatin (20 mg once a day) for the first 14 days of the study. Nelfinavir (1,250 mg twice a day) was added on days 15 to 28. Pharmacokinetic assessment was performed on days 14 and 28. The study drugs were well tolerated. Nelfinavir increased the steady-state area under the plasma concentration-time curve during one dosing period (AUC
τ
) of atorvastatin 74% and the maximum concentration (
C
max
) of atorvastatin 122% and increased the AUC
τ
of simvastatin 505% and the
C
max
of simvastatin 517%. Neither atorvastatin nor simvastatin appeared to alter the pharmacokinetics of nelfinavir. It is recommended that coadministration of simvastatin with nelfinavir should be avoided, whereas atorvastatin should be used with nelfinavir with caution.
Effects of Single Oral Doses of Gemifloxacin (320 Milligrams) versus Trovafloxacin (200 Milligrams) on Fecal Flora in Healthy Volunteers
Authors: Garcia-Calvo Gloria , Molleja Ana , Giménez Maria J. , Parra Araceli , Nieto Eva , Ponte Carmen , Aguilar Lorenzo , Soriano Francisco 
Published: July 2002

ABSTRACT

Gemifloxacin and trovafloxacin were administered to 12 volunteers in a randomized crossover trial with a 2-week washout period. Stool samples were collected predose and 1, 2, and 3 days postdose. Both quinolones reduced the number of organisms of the family
Enterobacteriaceae
and aerobic gram-positive organisms.
Escherichia coli
reduction was greater with gemifloxacin than with trovafloxacin, with postdose isolation of quinolone-resistant strains for which MICs of trovafloxacin were higher than those of gemifloxacin.