bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evidence for Active Efflux as the Primary Mechanism of Resistance to Ciprofloxacin in
Salmonella enterica
Serovar Typhimurium
Authors: Giraud Etienne , Cloeckaert Axel , Kerboeuf Dominique , Chaslus-Dancla Elisabeth 
Published: July 2002

ABSTRACT

The occurrence of active efflux and cell wall modifications were studied in
Salmonella enterica
serovar Typhimurium mutants that were selected with enrofloxacin and whose phenotypes of resistance to fluoroquinolones could not be explained only by mutations in the genes coding for gyrase or topoisomerase IV. Mutant BN18/21 exhibited a decreased susceptibility to ciprofloxacin (MIC = 0.125 μg/ml) but did not have a mutation in the
gyrA
gene. Mutants BN18/41 and BN18/71 had the same substitution, Gly81Cys in GyrA, but exhibited different levels of resistance to ciprofloxacin (MICs = 2 and 8 μg/ml, respectively). None of the mutants had mutations in the
parC
gene. Evidence for active efflux was provided by a classical fluorimetric method, which revealed a three- to fourfold decrease in ciprofloxacin accumulation in the three mutants compared to that in the parent strain, which was annuled by addition of the efflux pump inhibitor carbonyl cyanide
m
-chlorophenylhydrazone. In mutant BN18/71, a second fluorimetric method also showed a 50% reduction in the level of accumulation of ethidium bromide, a known efflux pump substrate. Immunoblotting and enzyme-linked immunosorbent assay experiments with an anti-AcrA antibody revealed that the resistance phenotype was strongly correlated with the expression level of the AcrAB efflux pump and suggested that decreased susceptibility to ciprofloxacin due to active efflux probably related to overproduction of this pump could occur before that due to
gyrA
mutations. Alterations were also found in the outer membrane protein and lipopolysaccharide profiles of the mutants, and these alterations were possibly responsible for the decrease in the permeability of the outer membrane that was observed in the mutants and that could act synergistically with active efflux to decrease the level of ciprofloxacin accumulation.
In Vitro Pharmacodynamics of the New Ketolides HMR 3004 and HMR 3647 (Telithromycin) against
Chlamydia pneumoniae
Authors: Gustafsson I. , Hjelm E. , Cars O. 
Published: July 2002

ABSTRACT

The ketolides HMR 3004 and HMR 3647 (telithromycin) are a new class of macrolides that have a potential clinical efficacy against intracellular pathogens. The objectives of this study were to investigate the MIC, minimum bactericidal concentration, and time-dependent killing of two
Chlamydia pneumoniae
strains of the two ketolides. The killing effect was also studied with a newly developed intracellular in vitro kinetic model. Furthermore, HMR 3647 was studied for the effect of a subinhibitory concentration of 0.5 times the MIC after a preexposure of 10 times the MIC during 12 h. The MICs for both strains were 0.0039 and 0.0156 mg/liter for HMR 3004 and HMR 3647, respectively. Killing with 10 times the MIC was time dependent, increasing from a 1-log-unit decrease in the number of inclusions per well at 48 h to a maximal effect of 2.8-log-unit decrease after 96 h. A preexposure of 10 times the MIC of HMR 3647 for 12 h followed by a subinhibitory concentration of 0.5 times the MIC increased the killing effect to a 1.2-log-unit reduction in inclusions per well. An exposure for 12 h gave poor reduction of inclusions, while a static dose of 10 times the MIC for 72 h showed a 2.2-log-unit reduction in inclusions per well. In the kinetic model, a small number of inclusions were detected after 72 h by one exposure of 10 times the MIC. Regrowth could not be detected after 120 h. The ketolides HMR 3004 and HMR 3647 have bactericidal activity and show a significant sub-MIC effect on the intracellular pathogen
C. pneumoniae
.
Efficacies of Moxifloxacin, Ciprofloxacin, and Vancomycin against Experimental Endocarditis Due to Methicillin-Resistant
Staphylococcus aureus
Expressing Various Degrees of Ciprofloxacin Resistance
Authors: Entenza J. M. , Que Y. A. , Vouillamoz J. , Glauser M. P. , Moreillon P. 
Published: July 2002

ABSTRACT

The new 8-methoxyquinolone moxifloxacin was tested against two ciprofloxacin-susceptible
Staphylococcus aureus
strains (strains P8 and COL) and two ciprofloxacin-resistant derivatives of strain P8 carrying a single
grlA
mutation (strain P8-4) and double
grlA
and
gyrA
mutations (strain P8-128). All strains were resistant to methicillin. The MICs of ciprofloxacin and moxifloxacin were 0.5 and 0.125 mg/liter, respectively, for P8; 0.25 and 0.125 mg/liter, respectively, for COL; 8 and 0.25 mg/liter, respectively, for P8-4; and ≥128 and 2 mg/liter, respectively, for P8-128. In vitro, the rate of spontaneous resistance of P8 and COL was 10
−7
on agar plates containing ciprofloxacin at two times the MIC, whereas it was ≤10
−10
on agar plates containing moxifloxacin at two times the MIC. Rats with experimental aortic endocarditis were treated with doses of drugs that simulate the kinetics in humans: moxifloxacin, 400 mg orally once a day; ciprofloxacin, 750 mg orally twice a day; or vancomycin, 1 g intravenously twice a day. Treatment was started either 12 or 24 h after infection and lasted for 3 days. Moxifloxacin treatment resulted in culture-negative vegetations in a total of 20 of 21 (95%) rats infected with P8, 10 of 11 (91%) rats infected with COL, and 19 of 24 (79%) rats infected with P8-4 (
P
< 0.05 compared to the results for the controls). In contrast, ciprofloxacin treatment sterilized zero of nine (0%) vegetations infected with first-level resistant mutant P8-4. Vancomycin sterilized only 8 of 15 (53%), 6 of 11 (54%), and 12 of 23 (52%) of the vegetations, respectively. No moxifloxacin-resistant derivative emerged among these organisms. However, moxifloxacin treatment of highly ciprofloxacin-resistant mutant P8-128 failed and selected for variants for which the MIC increased two times in 2 of 10 animals. Thus, while oral moxifloxacin might deserve consideration as treatment for staphylococcal infections in humans, caution related to its use against strains for which MICs are borderline is warranted.
In Vitro Activity of PNU-100766 (Linezolid), a New Oxazolidinone Antimicrobial, against
Nocardia brasiliensis
Authors: Vera-Cabrera Lucio , Gómez-Flores Alejandra , Escalante-Fuentes Wendy G. , Welsh Oliverio 
Published: July 2002

ABSTRACT

The in vitro activity of a novel oxazolidinone, linezolid, was studied by comparing the activity of linezolid with those of amikacin, trimethoprim-sulfamethoxazole, and amoxicillin-clavulanic acid against 25 strains of
Nocardia brasiliensis
isolated from patients with mycetoma. All
N. brasiliensis
strains tested were sensitive to linezolid (MIC at which 90% of strains are inhibited [MIC
90
], 2 μg/ml; MIC
50
, 1 μg/ml). This antimicrobial might constitute a good alternative for treatment of actinomycetoma.
Susceptibility Testing of
Aspergillus flavus
: Inoculum Dependence with Itraconazole and Lack of Correlation between Susceptibility to Amphotericin B In Vitro and Outcome In Vivo
Authors: Mosquera J. , Warn P. A. , Morrissey J. , Moore C. B. , Gil-Lamaignere C. , Denning D. W. 
Published: July 2002

ABSTRACT

We have attempted to validate in
Aspergillus flavus
the main in vitro methodologies that have been used to detect resistance in
Aspergillus fumigatus
. We developed a murine model with two
A. flavus
isolates, one that was apparently resistant in vitro to amphotericin B (AFL5) and another that was resistant to itraconazole (AFL8). No correlation was found for amphotericin B in AFL5, since the in vivo response was compatible with a susceptible isolate. Modification of the in vitro susceptibility test methodology for amphotericin B was unsuccessful. Although AFL8 was apparently resistant to itraconazole in vitro, it was found to be susceptible in vivo. Additional in vitro work has detected weaknesses in the in vitro susceptibility methodology validated for
A. fumigatus
when applied to
A. flavus
. The principal problems are that changes in the inoculum have a large effect on the MICs of itraconazole for some
A. flavus
strains and that a trailing end point and spore sediment often appear when an inoculum with a higher colony count is used. We propose a modified method using a final inoculum of 2.5 × 10
4
CFU per ml of RPMI 1640 medium with 2% glucose buffered to pH 7.0 in a microtiter format, incubated for 48 h with no growth end point. Validation of this methodology requires one or more itraconazole-resistant
A. flavus
isolates, which have yet to be identified.
RFI-641, a Potent Respiratory Syncytial Virus Inhibitor
Authors: Huntley Clayton C. , Weiss William J. , Gazumyan Anna , Buklan Aron , Feld Boris , Hu William , Jones Thomas R. , Murphy Timothy , Nikitenko Antonia A. , O'Hara Bryan , Prince Gregory , Quartuccio Susan , Raifeld Yuri E. , Wyde Philip , O'Connell John F. 
Published: July 2002

ABSTRACT
Human respiratory syncytial virus (RSV), a paramyxovirus, is a major cause of acute upper and lower respiratory tract infections in infants, young children, and adults. RFI-641 is a novel anti-RSV agent with potent in vitro and in vivo activity. RFI-641 is active against both RSV type A and B strains. The viral specificity and the large therapeutic window of RFI-641 (>100-fold) indicate that the antiviral activity of the compound is not due to adverse effects on normal cells. The potent in vitro activity of RFI-641 can be translated to efficacy in vivo: RFI-641 is efficacious when administered prophylactically by the intranasal route in mice, cotton rats, and African green monkeys. RFI-641 is also efficacious when administered therapeutically (24 h postinfection) in the monkey model. Mechanism of action studies indicate that RFI-641 blocks viral F protein-mediated fusion and cell syncytium formation.
Survival and Epiphytic Fitness of a Nonpathogenic Mutant of
Xanthomonas campestris
pv. Glycines
Authors: Rukayadi Yaya , Suwanto Antonius , Tjahjono Budi , Harling Rob 
Published: July 2002

ABSTRACT

Xanthomonas campestris
pv. glycines is the causal agent of bacterial pustule disease of soybeans. The objective of this work was to construct a nonpathogenic mutant derived from the pathogenic wild-type strain YR32 and to evaluate its effectiveness in preventing growth of its parent on the soybean phyllosphere. A mini-Tn
5
-derived transposon was used to generate nonpathogenic mutants. Southern hybridization and pulsed-field gel electrophoresis confirmed the presence of a single transposon in each of the nonpathogenic mutants. One of the nonpathogenic mutants, M715, failed to induce a hypersensitive response in tomato leaves. An ice nucleation gene (
inaZ
) carried in pJL1703 was introduced into strain YR32 as a reporter gene to demonstrate that the presence of M715 could reduce colonization of the soybean phyllosphere by YR32. de Wit serial replacement analysis showed that M715 competed equally with its wild-type parental strain, YR32. Epiphytic fitness analysis of YR32 in the greenhouse indicated that the population dynamics of strains YR32, YR32(pJL1703), and M715 were similar, although the density of the mutant was slightly less than that of its parent. The M715 mutant was able to survive for 16 days after inoculation on soybean leaves and maintained population densities of approximately 10
4
to 10
5
cells g (fresh weight) of leaf
−1
. Therefore, M715 shows promise as an effective biocontrol agent for bacterial pustule disease in soybeans.
An α-Proteobacterium Converts Linear Alkylbenzenesulfonate Surfactants into Sulfophenylcarboxylates and Linear Alkyldiphenyletherdisulfonate Surfactants into Sulfodiphenylethercarboxylates
Authors: Schleheck David , Dong Wenbo , Denger Karin , Heinzle Elmar , Cook Alasdair M. 
Published: July 2002

ABSTRACTThe surfactant linear alkylbenzenesulfonate (LAS; 0.5 mM) or linear monoalkyldiphenyletherdisulfonate (LADPEDS; 0.5 mM) in salts medium was easily degraded in laboratory trickling filters, whereas carbon-limited, aerobic enrichment cultures in suspended culture with the same inocula did not grow. We took portions of the trickling filters which degraded LADPEDS, shook the organisms from the solid support (polyester), and found that growth in suspended culture in LADPEDS-salts medium occurred only in the presence of some solid support (polyester fleece or glass wool), though little biomass was immobilized on the support. The end products in suspended culture were identical with those from the trickling filters. There was low plating efficiency of LADPEDS-grown cultures on complex medium, and no picked colony or mixture of colonies grew in LADPEDS-salts-glass wool medium. However, selective plates containing LADPEDS-salts medium solidified with agarose yielded LADPEDS-dependent, pinpoint colonies which could be picked singly and subcultured in selective liquid medium. Isolate DS-1 was a bacterium which showed 93% sequence homology (16S ribosomal DNA) to its nearest phylogenetic neighbor, an α-proteobacterium. Strain DS-1 grew heterotrophically in LADPEDS-salts-glass wool medium and converted the set of aryl-substituted alkanes to the corresponding aryl-substituted carboxylic acids of shorter chain length. Similarly, strain DS-1 grew heterotrophically with commercial LAS, converting it to a set of sulfophenylcarboxylates. Growth with a single isomer of LAS [3-(4-sulfophenyl)dodecane] was concomitant with excretion of 4-(4-sulfophenyl)hexanoate, which was identified by matrix-assisted laser desorption ionization mass spectrometry. The growth yield (6.4 g of protein/mol of C) indicated mass balance, which, with the specific growth rate (0.05 h−1), indicated a specific utilization rate of LAS of 2.2 mkat/kg of protein.
Phylogenetic Relationships of
Cryptosporidium
Parasites Based on the 70-Kilodalton Heat Shock Protein (HSP70) Gene
Authors: Sulaiman Irshad M. , Morgan Una M. , Thompson R. C. Andrew , Lal Altaf A. , Xiao Lihua 
Published: July 2002

ABSTRACT

We have characterized the nucleotide sequences of the 70-kDa heat shock protein (HSP70) genes of
Cryptosporidium baileyi
,
C. felis
,
C. meleagridis
,
C. muris
,
C. serpentis
,
C. wrairi
, and
C. parvum
from various animals. Results of the phylogenetic analysis revealed the presence of several genetically distinct species in the genus
Cryptosporidium
and eight distinct genotypes within the species
C. parvum
. Some of the latter may represent cryptic species. The phylogenetic tree constructed from these sequences is in agreement with our previous results based on the small-subunit rRNA genes of
Cryptosporidium
parasites. The
Cryptosporidium
species formed two major clades: isolates of
C. muris
and
C. serpentis
formed the first major group, while isolates of
C. felis
,
C. meleagridis
,
C. wrairi
, and eight genotypes of
C. parvum
formed the second major group. Sequence variations were also observed between
C. muris
isolates from ruminants and rodents. The HSP70 gene provides another useful locus for phylogenetic analysis of the genus
Cryptosporidium
.
Cyt1A from
Bacillus thuringiensis
Lacks Toxicity to Susceptible and Resistant Larvae of Diamondback Moth (
Plutella xylostella
) and Pink Bollworm (
Pectinophora gossypiella
)
Authors: Meyer Susan K. , Tabashnik Bruce E. , Liu Yong-Biao , Wirth Margaret C. , Federici Brian A. 
Published: July 2002

ABSTRACT

We tested Cyt1Aa, a cytolytic endotoxin of
Bacillus thuringiensis
, against susceptible and Cry1A-resistant larvae of two lepidopteran pests, diamondback moth (
Plutella xylostella
) and pink bollworm (
Pectinophora gossypiella
). Unlike previous results obtained with mosquito and beetle larvae, Cyt1Aa alone or in combination with Cry toxins was not highly toxic to the lepidopteran larvae that we examined.
Differential Effects of pH on the Pore-Forming Properties of
Bacillus thuringiensis
Insecticidal Crystal Toxins
Authors: Tran Le Binh , Vachon Vincent , Schwartz Jean-Louis , Laprade Raynald 
Published: July 2002

ABSTRACT

The effect of pH on the pore-forming ability of two
Bacillus thuringiensis
toxins, Cry1Ac and Cry1C, was examined with midgut brush border membrane vesicles isolated from the tobacco hornworm,
Manduca sexta
, and a light-scattering assay. In the presence of Cry1Ac, membrane permeability remained high over the entire pH range tested (6.5 to 10.5) for KCl and tetramethylammonium chloride, but was much lower at pH 6.5 than at higher pHs for potassium gluconate, sucrose, and raffinose. On the other hand, the Cry1C-induced permeability to all substrates tested was much higher at pH 6.5, 7.5, and 8.5 than at pH 9.5 and 10.5. These results indicate that the pores formed by Cry1Ac are significantly smaller at pH 6.5 than under alkaline conditions, whereas the pore-forming ability of Cry1C decreases sharply above pH 8.5. The reduced activity of Cry1C at high pH correlates well with the fact that its toxicity for
M. sexta
is considerably weaker than that of Cry1Aa, Cry1Ab, and Cry1Ac. However, Cry1E, despite having a toxicity comparable to that of Cry1C, formed channels as efficiently as the Cry1A toxins at pH 10.5. These results strongly suggest that although pH can influence toxin activity, additional factors also modulate toxin potency in the insect midgut.
Molecular Diversity of
Lactobacillus
spp. and Other Lactic Acid Bacteria in the Human Intestine as Determined by Specific Amplification of 16S Ribosomal DNA
Authors: Heilig Hans G.H.J. , Zoetendal Erwin G. , Vaughan Elaine E. , Marteau Philippe , Akkermans Antoon D.L. , de Vos Willem M. 
Published: July 2002

ABSTRACT

A
Lactobacillus
group-specific PCR primer, S-G-Lab-0677-a-A-17, was developed to selectively amplify 16S ribosomal DNA (rDNA) from lactobacilli and related lactic acid bacteria, including members of the genera
Leuconostoc
,
Pediococcus
, and
Weissella.
Amplicons generated by PCR from a variety of gastrointestinal (GI) tract samples, including those originating from feces and cecum, resulted predominantly in
Lactobacillus
-like sequences, of which ca. 28% were most similar to the 16S rDNA of
Lactobacillus ruminis
. Moreover, four sequences of
Leuconostoc
species were retrieved that, so far, have only been detected in environments other than the GI tract, such as fermented food products. The validity of the primer was further demonstrated by using
Lactobacillus
-specific PCR and denaturing gradient gel electrophoresis (DGGE) of the 16S rDNA amplicons of fecal and cecal origin from different age groups. The stability of the GI-tract bacterial community in different age groups over various time periods was studied. The
Lactobacillus
community in three adults over a 2-year period showed variation in composition and stability depending on the individual, while successional change of the
Lactobacillus
community was observed during the first 5 months of an infant’s life. Furthermore, the specific PCR and DGGE approach was tested to study the retention in fecal samples of a
Lactobacillus
strain administered during a clinical trial. In conclusion, the combination of specific PCR and DGGE analysis of 16S rDNA amplicons allows the diversity of important groups of bacteria that are present in low numbers in specific ecosystems to be characterized, such as the lactobacilli in the human GI tract.
Identification of a Gene Essential for Sheathed Structure Formation in
Sphaerotilus natans
, a Filamentous Sheathed Bacterium
Authors: Suzuki Toshihiko , Kanagawa Takahiro , Kamagata Yoichi 
Published: July 2002

ABSTRACT

Sphaerotilus natans
, a filamentous bacterium that causes bulking in activated sludge processes, can assume two distinct morphologies, depending on the substrate concentration for growth; in substrate-rich media it grows as single rod-shaped cells, whereas in substrate-limited media it grows as filaments. To identify genes responsible for sheath formation, we carried out transposon Tn
5
mutagenesis. Of the approximately 20,000 mutants obtained, 7 did not form sheathed structures. Sequencing of the Tn
5
-flanking regions showed that five of the seven Tn
5
insertions converged at the same open reading frame, designated
sthA
. The deduced amino acids encoded by
sthA
were found to be homologous to glycosyltransferase, which is known to be involved in linking sugars to lipid carriers during bacterial exopolysaccharide biosynthesis. Disruption of the gene of the wild-type strain by inserting a kanamycin resistance gene cassette also resulted in sheathless growth under either type of nutrient condition. These findings indicate that
sthA
is a crucial component responsible for sheath formation.
Calcium Phosphate Nanoparticle Adjuvant
Authors: He Qing , Mitchell Alaina R. , Johnson Stacy L. , Wagner-Bartak Claus , Morcol Tulin , Bell Steve J. D. 
Published: July 2002

ABSTRACT
Vaccination to protect against human infectious diseases may be enhanced by using adjuvants that can selectively stimulate immunoregulatory responses. In a murine model, a novel nanoparticulate adjuvant composed of calcium phosphate (CAP) was compared with the commonly used aluminum (alum) adjuvants for its ability to induce immunity to herpes simplex virus type 2 (HSV-2) and Epstein-Barr virus (EBV) infections. Results indicated that CAP was more potent as an adjuvant than alum, elicited little or no inflammation at the site of administration, induced high titers of immunoglobulin G2a (IgG2a) antibody and neutralizing antibody, and facilitated a high percentage of protection against HSV-2 infection. Additional benefits of CAP include (i) an insignificant IgE response, which is an important advantage over injection of alum compounds, and (ii) the fact that CAP is a natural constituent of the human body. Thus, CAP is very well tolerated and absorbed. These studies were performed with animal models. By virtue of the potency of this CAP adjuvant and the relative absence of side effects, we believe that this new CAP formulation has great potential for use as an adjuvant in humans.
DNA Sequence and Comparison of Virulence Plasmids from
Rhodococcus equi
ATCC 33701 and 103
Authors: Takai Shinji , Hines Stephen A. , Sekizaki Tsutomu , Nicholson Vivian M. , Alperin Debra A. , Osaki Makoto , Takamatsu Daisuke , Nakamura Mutsu , Suzuki Kayo , Ogino Nobuko , Kakuda Tsutomu , Dan Hanhong , Prescott John F. 
Published: July 2002

ABSTRACT

The virulence plasmids of the equine virulent strains
Rhodococcus equi
ATCC 33701 and 103 were sequenced, and their genetic structure was analyzed. p33701 was 80,610 bp in length, and p103 was 1 bp shorter; their sequences were virtually identical. The plasmids contained 64 open reading frames (ORFs), 22 of which were homologous with genes of known function and 3 of which were homologous with putative genes of unknown function in other species. Putative functions were assigned to five ORFs based on protein family characteristics. The most striking feature of the virulence plasmids was the presence of a 27,536-bp pathogenicity island containing seven virulence-associated protein (
vap
) genes, including
vapA
. These
vap
genes have extensive homology to
vapA
, which encodes a thermoregulated and surface-expressed protein. The pathogenicity island contained a LysR family transcriptional regulator and a two-component response regulator upstream of six of the
vap
genes. The
vap
genes were present as a cluster of three (
vapA
,
vapC
, and
vapD
), as a pair (
vapE
and
vapF
), or individually (
vapG
;
vapH
). A region of extensive direct repeats of unknown function, possibly associated with thermoregulation, was present immediately upstream of the clustered and the paired genes but not the individual
vap
genes. There was extensive homology among the C-terminal halves of all
vap
genes but not generally among the N-terminal halves. The remainder of the plasmid consisted of a large region which appears to be associated with conjugation functions and a large region which appears to be associated with replication and partitioning functions.