bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective Targeting of Topoisomerase IV and DNA Gyrase in
Staphylococcus aureus
: Different Patterns of Quinolone- Induced Inhibition of DNA Synthesis
Authors: Fournier Bénédicte , Zhao Xilin , Lu Tao , Drlica Karl , Hooper David C. 
Published: July 2002

ABSTRACT

The effect of quinolones on the inhibition of DNA synthesis in
Staphylococcus aureus
was examined by using single resistance mutations in
parC
or
gyrA
to distinguish action against gyrase or topoisomerase IV, respectively. Norfloxacin preferentially attacked topoisomerase IV and blocked DNA synthesis slowly, while nalidixic acid targeted gyrase and inhibited replication rapidly. Ciprofloxacin exhibited an intermediate response, consistent with both enzymes being targeted. The absence of RecA had little influence on target choice by this assay, indicating that differences in rebound (repair) DNA synthesis were not responsible for the results. At saturating drug concentrations, norfloxacin and a
gyrA
mutant were used to show that topoisomerase IV-norfloxacin-cleaved DNA complexes are distributed on the
S. aureus
chromosome at intervals of about 30 kbp. If cleaved complexes block DNA replication, as indicated by previous work, such close spacing of topoisomerase-quinolone-DNA complexes should block replication rapidly (replication forks are likely to encounter a cleaved complex within a minute). Thus, the slow inhibition of DNA synthesis at growth-inhibitory concentrations suggests that a subset of more distantly distributed complexes is physiologically relevant for drug action and is unlikely to be located immediately in front of the DNA replication fork.
Biochemical-Genetic Characterization and Distribution of OXA-22, a Chromosomal and Inducible Class D β-Lactamase fromRalstonia(Pseudomonas)pickettii
Authors: Nordmann Patrice , Poirel Laurent , Kubina Maryline , Casetta Anne , Naas Thierry 
Published: July 2002

ABSTRACTFrom genomic DNA ofRalstonia pickettiiisolate PIC-1, a β-lactamase gene was cloned that encodes the oxacillinase OXA-22. It differs from known oxacillinases, being most closely related to OXA-9 (38% amino acid identity). The hydrolytic spectrum of OXA-22 is limited mostly to benzylpenicillin, cloxacillin, and restricted-spectrum cephalosporins. OXA-22-like genes were identified as single chromosomal copies in five otherR. pickettiiclinical isolates. The expression of OXA-22-like β-lactamases was inducible inR. pickettii.
SmeDEF Multidrug Efflux Pump Contributes to Intrinsic Multidrug Resistance in
Stenotrophomonas maltophilia
Authors: Zhang Li , Li Xian-Zhi , Poole Keith 
Published: July 2002

ABSTRACT

Stenotrophomonas maltophilia
is an emerging nosocomial pathogen that displays high-level intrinsic resistance to a variety of structurally unrelated antimicrobial agents. Efflux mechanisms are known to contribute to acquired multidrug resistance in this organism, and indeed, one such multidrug efflux system, SmeDEF, was recently identified. Still, the importance of SmeDEF to intrinsic antibiotic resistance in
S. maltophilia
had not yet been determined. Reverse transcription-PCR confirmed expression of the
smeDEF
genes in wild-type
S. maltophilia
, and deletion of
smeE
or
smeF
in wild-type strains rendered the mutants hypersusceptible to several antimicrobials, suggesting that SmeDEF contributes to intrinsic antimicrobial resistance in this organism. Expression of
smeDEF
was also enhanced in an in vitro-selected multidrug-resistant mutant, although deletion of
smeF
but not of
smeE
in these mutants compromised antimicrobial resistance. Apparently, hyperexpressed SmeF is capable of functioning with additional multidrug efflux components to promote multidrug resistance in
S. maltophilia
.
Target Preference of 15 Quinolones againstStaphylococcus aureus, Based on Antibacterial Activities and Target Inhibition
Authors: Takei Masaya , Fukuda Hideyuki , Kishii Ryuta , Hosaka Masaki 
Published: July 2002

ABSTRACTThe antibacterial activities and target inhibition of 15 quinolones againstgrlAandgyrAmutant strains were studied. The strains were obtained from wild-typeStaphylococcus aureusMS5935 by selection with norfloxacin and nadifloxacin, respectively. The antibacterial activities of most quinolones against both mutant strains were lower than those against the wild-type strain. The ratios of MICs for thegyrAmutant strain to those for thegrlAmutant strain (MIC ratio) varied from 0.125 to 4. The ratios of 50% inhibitory concentrations (IC50s) of quinolones against topoisomerase IV to those against DNA gyrase (IC50ratios) also varied, from 0.177 to 5.52. A significant correlation between the MIC ratios and the IC50ratios was observed (r= 0.919;P< 0.001). These results suggest that the antibacterial activities of quinolones against the wild-type strain are involved not only in topoisomerase IV inhibition but also in DNA gyrase inhibition and that the target preference in the wild-type strain can be anticipated by the MIC ratios. Based on the MIC ratios, the quinolones were classified into three categories. Type I quinolones (norfloxacin, enoxacin, fleroxacin, ciprofloxacin, lomefloxacin, trovafloxacin, grepafloxacin, ofloxacin, and levofloxacin) had MIC ratios of <1, type II quinolones (sparfloxacin and nadifloxacin) had MIC ratios of >1, and type III quinolones (gatifloxacin, pazufloxacin, moxifloxacin, and clinafloxacin) had MIC ratios of 1. Type I and type II quinolones seem to prefer topoisomerase IV and DNA gyrase, respectively. Type III quinolones seem to target both enzymes at nearly the same level in bacterial cells (a phenomenon known as the dual-targeting property), and their IC50ratios were approximately 2.
Need for Annual Surveillance of Antimicrobial Resistance in
Streptococcus pneumoniae
in the United States: 2-Year Longitudinal Analysis
Authors: Sahm Daniel F. , Karlowsky James A. , Kelly Laurie J. , Critchley Ian A. , Jones Mark E. , Thornsberry Clyde , Mauriz Yolanda , Kahn James 
Published: July 2002

ABSTRACT

Although changing patterns in antimicrobial resistance in
Streptococcus pneumoniae
have prompted several surveillance initiatives in recent years, the frequency with which these studies are needed has not been addressed. To approach this issue, the extent to which resistance patterns change over a 1-year period was examined. In this study we analyzed
S. pneumoniae
antimicrobial susceptibility results produced in our laboratory with isolates obtained over 2 consecutive years (1997–1998 and 1998–1999) from the same 96 institutions distributed throughout the United States. Comparison of results revealed increases in resistant percentages for all antimicrobial agents studied except vancomycin. For four of the agents tested (penicillin, cefuroxime, trimethoprim-sulfamethoxazole, and levofloxacin), the increases were statistically significant (
P
< 0.05). Resistance to the fluoroquinolone remained low in both years (0.1 and 0.6%, respectively); in contrast, resistance to macrolides was consistently greater than 20%, and resistance to trimethoprim-sulfamethoxazole increased from 13.3 to 27.3%. Multidrug resistance, concurrent resistance to three or more antimicrobials of different chemical classes, also increased significantly between years, from 5.9 to 11%. The most prevalent phenotype was resistance to penicillin, azithromycin (representative macrolide), and trimethoprim-sulfamethoxazole. Multidrug-resistant phenotypes that included fluoroquinolone resistance were uncommon; however, two phenotypes that included fluoroquinolone resistance not found in 1997–1998 were encountered in 1998–1999. This longitudinal surveillance study of resistance in
S. pneumoniae
revealed that significant changes do occur in just a single year and supports the need for surveillance at least on an annual basis, if not continuously.
Staphylococcus aureus
Cell Extract Transcription-Translation Assay: Firefly Luciferase Reporter System for Evaluating Protein Translation Inhibitors
Authors: Murray Robert W. , Melchior Earline P. , Hagadorn Jeanne C. , Marotti Keith R. 
Published: July 2002

ABSTRACT

The promoter for the
Staphylococcus aureus
capsule polysaccharide synthesis gene (
cap1A
) was cloned in front of the firefly luciferase gene for use in a cell extract
S. aureus
transcription-translation system. The assay is rapid, reproducible, and sensitive and has a lower background level than the radiolabeled amino acid incorporation translation assays. We present data evaluating a transcription inhibitor and a number of protein translation inhibitors in this system.
SmeC, an Outer Membrane Multidrug Efflux Protein of
Stenotrophomonas maltophilia
Authors: Li Xian-Zhi , Zhang Li , Poole Keith 
Published: July 2002

ABSTRACT

A homologue of the
mexAB-oprM
multidrug efflux operon of
Pseudomonas aeruginosa
,
smeABC
, was cloned from
Stenotrophomonas maltophilia
by using, as a probe, a PCR product amplified from this organism with primers based on the
mexB
sequence. The
smeABC
genes were hyperexpressed in a mutant strain displaying resistance to several antimicrobials, including aminoglycosides, β-lactams, and fluoroquinolones. Deletions in
smeC
but not
smeB
compromised this resistance, suggesting that SmeC contributed to the multidrug resistance of the mutant as part of another, as-yet-unidentified multidrug efflux system. Consistent with SmeC functioning independently of SmeAB, a promoter activity was identified upstream of
smeC
. Upstream of the
smeABC
genes, a putative two-gene operon,
smeSR
, encoding homologues of bacterial two-component regulatory systems was identified. The cloned
smeR
gene activated expression of a
smeA-lacZ
fusion, indicating that SmeR positively regulates expression of the
smeABC
genes. Consistent with this, the multidrug resistance of the SmeABC-hyperexpressing mutant was compromised by deletion of
smeR
. Intriguingly, SmeC expression in
S. maltophilia
paralleled a β-lactamase activity provided by a C-terminally truncated L2 enzyme, which was apparently responsible for the β-lactam resistance of the SmeABC-hyperexpressing mutant. This represents the first report of coregulation of an efflux resistance determinant and a β-lactamase.
Resistance Mechanisms in Clinical Isolates of
Candida albicans
Authors: White Theodore C. , Holleman Scott , Dy Francis , Mirels Laurence F. , Stevens David A. 
Published: July 2002

ABSTRACT

Resistance to azole antifungals continues to be a significant problem in the common fungal pathogen
Candida albicans
. Many of the molecular mechanisms of resistance have been defined with matched sets of susceptible and resistant clinical isolates from the same strain. Mechanisms that have been identified include alterations in the gene encoding the target enzyme
ERG11
or overexpression of efflux pump genes including
CDR1
,
CDR2
, and
MDR1
. In the present study, a collection of unmatched clinical isolates of
C. albicans
was analyzed for the known molecular mechanisms of resistance by standard methods. The collection was assembled so that approximately half of the isolates were resistant to azole drugs. Extensive cross-resistance was observed for fluconazole, clotrimazole, itraconazole, and ketoconazole. Northern blotting analyses indicated that overexpression of
CDR1
and
CDR2
correlates with resistance, suggesting that the two genes may be coregulated.
MDR1
overexpression was observed infrequently in some resistant isolates. Overexpression of
FLU1
, an efflux pump gene related to
MDR1
, did not correlate with resistance, nor did overexpression of
ERG11
. Limited analysis of the
ERG11
gene sequence identified several point mutations in resistant isolates; these mutations have been described previously. Two of the most common point mutations in
ERG11
associated with resistance, D116E and E266D, were tested by restriction fragment length polymorphism analysis of the isolates from this collection. The results indicated that the two mutations occur frequently in different isolates of
C. albicans
and are not reliably associated with resistance. These analyses emphasize the diversity of mechanisms that result in a phenotype of azole resistance. They suggest that the resistance mechanisms identified in matched sets of susceptible and resistant isolates are not sufficient to explain resistance in a collection of unmatched clinical isolates and that additional mechanisms have yet to be discovered.
Helicobacter pylori
Primary Resistance to Metronidazole and Clarithromycin in Brazil
Authors: Prazeres Magalhães Paula , de Magalhães Queiroz Dulciene Maria , Campos Barbosa Daniela Vale , Aguiar Rocha Gifone , Nogueira Mendes Edilberto , Santos Adriana , Valle Corrêa Paulo Renato , Camargos Rocha Andreia Maria , Martins Teixeira Lúcia , Affonso de Oliveira Celso 
Published: July 2002

ABSTRACT

Helicobacter pylori
resistance to metronidazole was detected in 107 (52.97%) of 202 strains. Twenty (9.85%) strains, 18 of them harboring 23S ribosomal DNA mutations, were resistant to clarithromycin. Metronidazole resistance was associated with female gender. Resistance to metronidazole and resistance to clarithromycin were associated. Increasing clarithromycin resistance rates were observed over time.
Bacterial Phosphating of Mild (Unalloyed) Steel
Authors: Volkland Hans-Peter , Harms Hauke , Müller Beat , Repphun Gernot , Wanner Oskar , Zehnder Alexander J. B. 
Published: July 2002

ABSTRACT

Mild (unalloyed) steel electrodes were incubated in phosphate-buffered cultures of aerobic, biofilm-forming
Rhodococcus
sp. strain C125 and
Pseudomonas putida
mt2. A resulting surface reaction leading to the formation of a corrosion-inhibiting vivianite layer was accompanied by a characteristic electrochemical potential (E) curve. First, E increased slightly due to the interaction of phosphate with the iron oxides covering the steel surface. Subsequently, E decreased rapidly and after 1 day reached −510 mV, the potential of free iron, indicating the removal of the iron oxides. At this point, only scattered patches of bacteria covered the surface. A surface reaction, in which iron was released and vivianite precipitated, started. E remained at −510 mV for about 2 days, during which the vivianite layer grew steadily. Thereafter, E increased markedly to the initial value, and the release of iron stopped. Changes in E and formation of vivianite were results of bacterial activity, with oxygen consumption by the biofilm being the driving force. These findings indicate that biofilms may protect steel surfaces and might be used as an alternative method to combat corrosion.
Identification of Species and Sources of
Cryptosporidium
Oocysts in Storm Waters with a Small-Subunit rRNA-Based Diagnostic and Genotyping Tool
Authors: Xiao Lihua , Alderisio Kerri , Limor Josef , Royer Michael , Lal Altaf A. 
Published: July 2002

ABSTRACT

The identification of
Cryptosporidium
oocysts in environmental samples is largely made by the use of an immunofluorescent assay. In this study, we have used a small-subunit rRNA-based PCR-restriction fragment length polymorphism technique to identify species and sources of
Cryptosporidium
oocysts present in 29 storm water samples collected from a stream in New York. A total of 12 genotypes were found in 27 positive samples; for 4 the species and probable origins were identified by sequence analysis, whereas the rest represent new genotypes from wildlife. Thus, this technique provides an alternative method for the detection and differentiation of
Cryptosporidium
parasites in environmental samples.
Identification of and Spatio-Temporal Differences between Microbial Assemblages from Two Neighboring Sulfurous Lakes: Comparison by Microscopy and Denaturing Gradient Gel Electrophoresis
Authors: Casamayor Emilio O. , Schäfer Hendrik , Bañeras Lluis , Pedrós-Alió Carlos , Muyzer Gerard 
Published: July 2002

ABSTRACT

The microbial assemblages of Lake Cisó and Lake Vilar (Banyoles, northeast Spain) were analyzed in space and time by microscopy and by performing PCR-denaturing gradient gel electrophoresis (DGGE) and sequence analysis of 16S rRNA gene fragments. Samples obtained from different water depths and at two different times of the year (in the winter during holomixis and in the early spring during a phytoplankton bloom) were analyzed. Although the lakes have the same climatic conditions and the same water source, the limnological parameters were different, as were most of the morphologically distinguishable photosynthetic bacteria enumerated by microscopy. The phylogenetic affiliations of the predominant DGGE bands were inferred by performing a comparative 16S rRNA sequence analysis. Sequences obtained from Lake Cisó samples were related to gram-positive bacteria and to members of the division
Proteobacteria
. Sequences obtained from Lake Vilar samples were related to members of the
Cytophaga-Flavobacterium-Bacteroides
phylum and to cyanobacteria. Thus, we found that like the previously reported differences between morphologically distinct inhabitants of the two lakes, there were also differences among the community members whose morphologies did not differ conspicuously. The changes in the species composition from winter to spring were also marked. The two lakes both contained sequences belonging to phototrophic green sulfur bacteria, which is consistent with microscopic observations, but these sequences were different from the sequences of cultured strains previously isolated from the lakes. Euryarchaeal sequences (i.e., methanogen- and thermoplasma-related sequences) also were present in both lakes. These euryarchaeal group sequences dominated the archaeal sequences in Lake Cisó but not in Lake Vilar. In Lake Vilar, a new planktonic population related to the crenarchaeota produced the dominant archaeal band. The phylogenetic analysis indicated that new bacterial and archaeal lineages were present and that the microbial diversity of these assemblages was greater than previously known. We evaluated the correspondence between the abundances of several morphotypes and DGGE bands by comparing microscopy and sequencing results. Our data provide evidence that the sequences obtained from the DGGE fingerprints correspond to the microorganisms that are actually present at higher concentrations in the natural system.
The Function of Cytoplasmic Flavin Reductases in the Reduction of Azo Dyes by Bacteria
Authors: Russ Rainer , Rau Jörg , Stolz Andreas 
Published: July 2002

ABSTRACT

A flavin reductase, which is naturally part of the ribonucleotide reductase complex of
Escherichia coli
, acted in cell extracts of recombinant
E. coli
strains under aerobic and anaerobic conditions as an “azo reductase.” The transfer of the recombinant plasmid, which resulted in the constitutive expression of high levels of activity of the flavin reductase, increased the reduction rate for different industrially relevant sulfonated azo dyes in vitro almost 100-fold. The flavin reductase gene (
fre
) was transferred to
Sphingomonas
sp. strain BN6, a bacterial strain able to degrade naphthalenesulfonates under aerobic conditions. The flavin reductase was also synthesized in significant amounts in the
Sphingomonas
strain. The reduction rates for the sulfonated azo compound amaranth were compared for whole cells and cell extracts from both recombinant strains,
E. coli
, and wild-type
Sphingomonas
sp. strain BN6. The whole cells showed less than 2% of the specific activities found with cell extracts. These results suggested that the cytoplasmic anaerobic “azo reductases,” which have been described repeatedly in in vitro systems, are presumably flavin reductases and that in vivo they have insignificant importance in the reduction of sulfonated azo compounds.
Identification of Nonpoint Sources of Fecal Pollution in Coastal Waters by Using Host-Specific 16S Ribosomal DNA Genetic Markers from Fecal Anaerobes
Authors: Bernhard Anne E. , Field Katharine G. 
Published: July 2002

ABSTRACT

We describe a new PCR-based method for distinguishing human and cow fecal contamination in coastal waters without culturing indicator organisms, and we show that the method can be used to track bacterial marker sequences in complex environments. We identified two human-specific genetic markers and five cow-specific genetic markers in fecal samples by amplifying 16S ribosomal DNA (rDNA) fragments from members of the genus
Bifidobacterium
and the
Bacteroides-Prevotella
group and performing length heterogeneity PCR and terminal restriction fragment length polymorphism analyses. Host-specific patterns suggested that there are species composition differences in the
Bifidobacterium
and
Bacteroides-Prevotella
populations of human and cow feces. The patterns were highly reproducible among different hosts belonging to the same species. Additionally, all host-specific genetic markers were detected in water samples collected from areas frequently contaminated with fecal pollution. Ease of detection and longer survival in water made
Bacteroides-Prevotella
indicators better than
Bifidobacterium
indicators. Fecal 16S rDNA sequences corresponding to our
Bacteroides-Prevotella
markers comprised closely related gene clusters, none of which exactly matched previously published
Bacteroides
or
Prevotella
sequences. Our method detected host-specific markers in water at pollutant concentrations of 2.8 × 10
−5
to 2.8 × 10
−7
g (dry weight) of feces/liter and 6.8 × 10
−7
g (dry weight) of sewage/liter. Although our aim was to identify nonpoint sources of fecal contamination, the method described here should be widely applicable for monitoring spatial and temporal fluctuations in specific bacterial groups in natural environments.
Expanding the Known Diversity and Environmental Distribution of an Uncultured Phylogenetic Division of Bacteria
Authors: Dojka Michael A. , Harris J. Kirk , Pace Norman R. 
Published: July 2002

ABSTRACT
Culture-independent molecular phylogenetic methods were used to explore the breadth of diversity and environmental distribution of members of the division-level “candidate” phylogenetic group WS6, recently discovered in a contaminated aquifer and with no cultivated representatives. A broad diversity of WS6-affiliated sequences were cloned from 7 of 12 environments investigated: mainly from anaerobic sediment environments. The number of sequences representing the WS6 candidate division was increased from 3 to 60 in this study. The extent of phylogenetic divergence (sequence difference) in this candidate division was found to be among the largest of any known bacterial division. This indicates that organisms representing the WS6 phylogenetic division offer a broad diversity of undiscovered biochemical and metabolic novelty. These results provide a framework for the further study of these evidently important kinds of organisms and tools, the sequences, with which to do so.