bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genotypic Analysis of
Mycobacterium tuberculosis
in Two Distinct Populations Using Molecular Beacons: Implications for Rapid Susceptibility Testing
Authors: Piatek Amy S. , Telenti Amalio , Murray Megan R. , El-Hajj Hiyam , Jacobs William R. , Kramer Fred Russell , Alland David 
Published: April 2009

ABSTRACT

Past genotypic studies of
Mycobacterium tuberculosis
may have incorrectly estimated the importance of specific drug resistance mutations due to a number of sampling biases including an overrepresentation of multidrug-resistant (MDR) isolates. An accurate assessment of resistance mutations is crucial for understanding basic resistance mechanisms and designing genotypic drug resistance assays. We developed a rapid closed-tube PCR assay using fluorogenic reporter molecules called molecular beacons to detect reportedly common
M. tuberculosis
mutations associated with resistance to isoniazid and rifampin. The assay was used in a comparative genotypic investigation of two different study populations to determine whether these known mutations account for most cases of clinical drug resistance. We analyzed samples from a reference laboratory in Madrid, Spain, which receives an overrepresentation of MDR isolates similar to prior studies and from a community medical center in New York where almost all of the resistant isolates and an equal number of susceptible controls were available. The ability of the molecular beacon assay to predict resistance to isoniazid and rifampin was also assessed. The overall sensitivity and specificity of the assay for isoniazid resistance were 85 and 100%, respectively, and those for rifampin resistance were 98 and 100%, respectively. Rifampin resistance mutations were detected equally well in isolates from both study populations; however, isoniazid resistance mutations were detected in 94% of the isolates from Madrid but in only 76% of the isolates from New York (
P
= 0.02). In New York, isoniazid resistance mutations were significantly more common in the MDR isolates (94%) than in single-drug-resistant isolates (44%;
P
< 0.001). No association between previously described mutations in the
kasA
gene and isoniazid resistance was found. The first mutations that cause isoniazid resistance may often occur in sequences that have not been commonly associated with isoniazid resistance, possibly in other as yet uncharacterized genes. The molecular beacon assay was simple, rapid, and highly sensitive for the detection of rifampin-resistant
M. tuberculosis
isolates and for the detection of isoniazid resistance in MDR isolates.
Development of a Long-Term Ascending Urinary Tract Infection Mouse Model for Antibiotic Treatment Studies
Authors: Hvidberg Hanne , Struve Carsten , Krogfelt Karen A. , Christensen Nils , Rasmussen Søren N. , Frimodt-Møller Niels 
Published: April 2009

ABSTRACT

A model of ascending unobstructed urinary tract infection (UTI) in mice was developed to study the significance of the antibiotic concentration in urine, serum, and kidney tissue for efficacy of treatment of UTI in general and pyelonephritis in particular. Outbred Ssc-CF1 female mice were used throughout the study, and
Escherichia coli
was used as the pathogen. The virulence of 11 uropathogenic
E. coli
isolates and 1 nonpathogenic laboratory
E. coli
strain was examined. Strain C175-94 achieved the highest counts in the kidneys, and this strain was subsequently used as the infecting organism. The model gave reproducible bladder infections, i.e., bacteria were recovered from 22 of 23 control mice after 3 days, and histological examination of kidney tissue showed that of 14 infected kidneys, 7 (50%) showed major histological changes, whereas 3 of 36 uninfected kidneys showed major histological changes (
P
= 0.018). Once the model was established, the efficacies of different doses of cefuroxime and gentamicin, corresponding to active concentrations in urine only or in urine, serum, and kidney tissue simultaneously, were examined. All cefuroxime doses resulted in significantly lower counts in urine than control treatments, but the dose which produced concentrations of cefuroxime only in urine and not in serum or kidney tissue had no effect on kidney infection. Even low doses of gentamicin (0.05 mg/mouse) resulted in concentrations in renal tissue for prolonged times due to accumulation. All gentamicin doses had a significant effect (compared to the effect of the control treatment) on bacterial counts in urine and kidneys. The antibiotic effect on bacterial counts in bladders was negligible for unknown reasons. Use of the mouse UTI model is feasible for study of the effect of an antibiotic in the urinary system, although the missing antibacterial effect in the bladder needs further evaluation.
In Vitro Susceptibilities of Rapidly Growing Mycobacteria to Telithromycin (HMR 3647) and Seven Other Antimicrobials
Authors: Fernández-Roblas Ricardo , Esteban Jaime , Cabria Froilán , López Juan Carlos , Jiménez Maria Soledad , Soriano Francisco 
Published: April 2009

ABSTRACT

The antimicrobial activities of telithromycin (HMR 3647) and seven other antimicrobials against 94 strains of rapidly growing mycobacteria were determined. Telithromycin at a concentration of 1 μg/ml inhibited
Mycobacterium peregrinum
(100%),
Mycobacterium chelonae
(80%),
Mycobacterium abscessus-Mycobacterium mucogenicum
(44.4%), and
Mycobacterium fortuitum
(2.1%). All or most strains of
M. peregrinum
,
M. fortuitum
, and
M. mucogenicum
were inhibited by 2 μg of quinolones per ml.
Morphological Change in
Pseudomonas aeruginosa
following Antibiotic Treatment of Experimental Infection in Mice and Its Relation to Susceptibility to Phagocytosis and to Release of Endotoxin
Authors: Yokochi Takashi , Narita Kayou , Morikawa Akiko , Takahashi Kazuko , Kato Yutaka , Sugiyama Tuyoshi , Koide Naoki , Kawai Makoto , Fukada Masako , Yoshida Tomoaki 
Published: April 2009

ABSTRACT

The relationship between morphological changes in
Pseudomonas aeruginosa
following antibiotic treatment of experimental infection in mice, susceptibility to phagocytosis, and release of endotoxin was studied. The intraperitoneal administration of
P. aeruginosa
with imipenem or ceftazidime into mice induced morphological changes in the cells 2 h after injection. Round
P. aeruginosa
cells with imipenem treatment became susceptible to phagocytosis by peritoneal cells, whereas long filamentous cells with ceftazidime treatment were hardly phagocytized by peritoneal cells. The morphological changes also affected the plasma endotoxin level in the circulation.
Production of a TEM-24 Plasmid-Mediated Extended-Spectrum β-Lactamase by a Clinical Isolate of
Pseudomonas aeruginosa
Authors: Marchandin Helene , Jean-Pierre Helene , De Champs Christophe , Sirot Danielle , Darbas Helene , Perigault Pierre Francois , Carriere Christian 
Published: April 2009

ABSTRACT

Several
Pseudomonas aeruginosa
strains, including one urinary isolate producing an extended-spectrum β-lactamase TEM-24, were isolated from a long-term-hospitalized woman. Three TEM-24-producing enterobacterial species (
Enterobacter aerogenes
,
Escherichia coli
, and
Proteus mirabilis
) were isolated from the same patient. TEM-24 and the resistance markers for aminoglycosides, chloramphenicol, and sulfonamide were encoded by a 180-kb plasmid transferred by conjugation into
E. coli
HB101.
In Vitro Activities of the New Antifungal Triazole SCH 56592 against Common and Emerging Yeast Pathogens
Authors: Barchiesi Francesco , Arzeni Daniela , Fothergill Annette W. , Di Francesco Luigi Falconi , Caselli Francesca , Rinaldi Michael G. , Scalise Giorgio 
Published: April 2009

ABSTRACT

A broth microdilution method performed in accordance with the National Committee for Clinical Laboratory Standards guidelines was used to compare the in vitro activity of the new antifungal triazole SCH 56592 (SCH) to that of fluconazole (FLC), itraconazole (ITC), and ketoconazole (KETO) against 257 clinical yeast isolates. They included 220 isolates belonging to 12 different species of
Candida
, 15 isolates each of
Cryptococcus neoformans
and
Saccharomyces cerevisiae
, and seven isolates of
Rhodotorula rubra
. The MICs of SCH at which 50% (MIC
50
) and 90% (MIC
90
) of the isolates were inhibited were 0.06 and 2.0 μg/ml, respectively. In general, SCH was considerably more active than FLC (MIC
50
and MIC
90
of 1.0 and 64 μg/ml, respectively) and slightly more active than either ITC (MIC
50
and MIC
90
of 0.25 and 2.0 μg/ml, respectively) and KETO (MIC
50
and MIC
90
of 0.125 and 4.0 μg/ml, respectively). Our in vitro data suggest that SCH has significant potential for clinical development.
Activities of Synthetic Hybrid Peptides against Anaerobic Bacteria: Aspects of Methodology and Stability
Authors: Oh Herin , Hedberg Maria , Wade David , Edlund Charlotta 
Published: April 2009

ABSTRACT

The increasing problem of antibiotic resistance among pathogenic bacteria requires development of new antimicrobial agents. One line of investigation is the synthesis of antimicrobial hybrid peptides. The aim of the present investigation was to determine the in vitro activities of 16 cecropin-melittin hybrid peptides (CAMEL analogues) against 60 anaerobic bacterial strains, to compare their activities with those of seven clinically used antimicrobial agents, and to compare different methods for anaerobic susceptibility testing of these peptides. The stability of one of the peptides, temporin B, with different stereoisomeric configurations was investigated in a fecal milieu. The CAMEL analogues showed antimicrobial activity against the anaerobic bacteria, with MICs ranging from 0.125 to 32 μg/ml. The overall activities (the MICs at which 90% of isolates are inhibited) of the CAMEL analogues against anaerobic bacteria were mainly inferior to those of imipenem, clindamycin, and piperacillin but were equal to or superior to those of metronidazole, cefoxitin, ciprofloxacin, and chloramphenicol. The agarose dilution method was found to be an accurate method for the testing of large numbers of bacterial strains. The
d
isomer of temporin B was inactivated more slowly in feces than the
l
isomer. This study shows that the CAMEL analogues are potential agents for the treatment of anaerobic infections.
Induction of a Futile Embden-Meyerhof-Parnas Pathway in
Deinococcus radiodurans
by Mn: Possible Role of the Pentose Phosphate Pathway in Cell Survival
Authors: Zhang Y.-M. , Wong T.-Y. , Chen L.-Y. , Lin C.-S. , Liu J.-K. 
Published: April 2009

ABSTRACT

Statistical models were used to predict the effects of tryptone, glucose, yeast extract (TGY) and Mn on biomass formation of the highly radioresistant bacterium
Deinococcus radiodurans
. Results suggested that glucose had marginal effect on biomass buildup, but Mn was a significant factor for biomass formation. Mn also facilitated glucose interactions with other nutrient components. These predictions were verified by in vivo and in vitro experiments. TGY-grown cells metabolized glucose solely by the pentose phosphate pathway (PPP). Although only a fraction of glucose from the medium was transported into the cells, glucose was incorporated into the DNA efficiently after cells were exposed to UV light. The presence of glucose also enhanced the radioresistance of the culture. Mn could induce an Embden-Meyerhof-Parnas (EMP) pathway in
D. radiodurans
. The EMP pathway and the PPP of the Mn-treated cells oxidized glucose simultaneously at a 6:1 ratio. Although glucose was hydrolyzed rapidly by the Mn-treated cells, most glucose was released as CO
2
. Mn-treated cultures retained less glucose per cell than cells grown without Mn, and still less glucose was incorporated into the DNA after cells were exposed to UV light. Mn-treated cells were also more sensitive to UV light. Results suggested that metabolites of glucose generated from the PPP enhanced the survival of
D. radiodurans
. Induction of the EMP pathway by Mn may deplete metabolites for DNA repair and may induce oxidative stress for the cell, leading to reduction of radioresistance.
Characterization of Two Inducible Phosphate Transport Systems in
Rhizobium tropici
Authors: Botero Lina M. , Al-Niemi Thamir S. , McDermott Timothy R. 
Published: April 2009

ABSTRACT

Rhizobium tropici
forms nitrogen-fixing nodules on the roots of the common bean (
Phaseolus vulgaris
). Like other legume-
Rhizobium
symbioses, the bean-
R. tropici
association is sensitive to the availability of phosphate (P
i
). To better understand phosphorus movement between the bacteroid and the host plant, P
i
transport was characterized in
R. tropici
. We observed two P
i
transport systems, a high-affinity system and a low-affinity system. To facilitate the study of these transport systems, a Tn
5
B22 transposon mutant lacking expression of the high-affinity transport system was isolated and used to characterize the low-affinity transport system in the absence of the high-affinity system. The

K
m

and
V
max
values for the low-affinity system were estimated to be 34 ± 3 μM P
i
and 118 ± 8 nmol of P
i
· min
−1
· mg (dry weight) of cells
−1
, respectively, and the

K
m

and
V
max
values for the high-affinity system were 0.45 ± 0.01 μM P
i
and 86 ± 5 nmol of P
i
· min
−1
· mg (dry weight) of cells
−1
, respectively. Both systems were inducible by P
i
starvation and were also shock sensitive, which indicated that there was a periplasmic binding-protein component. Neither transport system appeared to be sensitive to the proton motive force dissipator carbonyl cyanide
m
-chlorophenylhydrazone, but P
i
transport through both systems was eliminated by the ATPase inhibitor
N
,
N
′-dicyclohexylcarbodiimide; the P
i
transport rate was correlated with the intracellular ATP concentration. Also, P
i
movement through both systems appeared to be unidirectional, as no efflux or exchange was observed with either the wild-type strain or the mutant. These properties suggest that both P
i
transport systems are ABC type systems. Analysis of the transposon insertion site revealed that the interrupted gene exhibited a high level of homology with
kdpE
, which in several bacteria encodes a cytoplasmic response regulator that governs responses to low potassium contents and/or changes in medium osmolarity.
Evidence for Microbial Fe(III) Reduction in Anoxic, Mining-Impacted Lake Sediments (Lake Coeur d'Alene, Idaho)
Authors: Cummings David E. , March Anthony W. , Bostick Benjamin , Spring Stefan , Caccavo Frank , Fendorf Scott , Rosenzweig R. Frank 
Published: April 2009

ABSTRACT

Mining-impacted sediments of Lake Coeur d'Alene, Idaho, contain more than 10% metals on a dry weight basis, approximately 80% of which is iron. Since iron (hydr)oxides adsorb toxic, ore-associated elements, such as arsenic, iron (hydr)oxide reduction may in part control the mobility and bioavailability of these elements. Geochemical and microbiological data were collected to examine the ecological role of dissimilatory Fe(III)-reducing bacteria in this habitat. The concentration of mild-acid-extractable Fe(II) increased with sediment depth up to 50 g kg
−1
, suggesting that iron reduction has occurred recently. The maximum concentrations of dissolved Fe(II) in interstitial water (41 mg liter
−1
) occurred 10 to 15 cm beneath the sediment-water interface, suggesting that sulfidogenesis may not be the predominant terminal electron-accepting process in this environment and that dissolved Fe(II) arises from biological reductive dissolution of iron (hydr)oxides. The concentration of sedimentary magnetite (Fe
3
O
4
), a common product of bacterial Fe(III) hydroxide reduction, was as much as 15.5 g kg
−1
. Most-probable-number enrichment cultures revealed that the mean density of Fe(III)-reducing bacteria was 8.3 × 10
5
cells g (dry weight) of sediment
−1
. Two new strains of dissimilatory Fe(III)-reducing bacteria were isolated from surface sediments. Collectively, the results of this study support the hypothesis that dissimilatory reduction of iron has been and continues to be an important biogeochemical process in the environment examined.
Purification and Partial Characterization of a Murein Hydrolase, Millericin B, Produced by
Streptococcus milleri
NMSCC 061
Authors: Beukes M. , Bierbaum G. , Sahl H.-G. , Hastings J. W. 
Published: April 2009

ABSTRACT

Streptococcus milleri
NMSCC 061 was screened for antimicrobial substances and shown to produce a bacteriolytic cell wall hydrolase, termed millericin B. The enzyme was purified to homogeneity by a four-step purification procedure that consisted of ammonium sulfate precipitation followed by gel filtration, ultrafiltration, and ion-exchange chromatography. The yield following ion-exchange chromatography was 6.4%, with a greater-than-2,000-fold increase in specific activity. The molecular weight of the enzyme was 28,924 as determined by electrospray mass spectrometry. The amino acid sequences of both the N terminus of the enzyme (NH
2
SENDFSLAMVSN) and an internal fragment which was generated by cyanogen bromide cleavage (NH
2
SIQTNAPWGL) were determined by automated Edman degradation. Millericin B displayed a broad spectrum of activity against gram-positive bacteria but was not active against
Bacillus subtilis
W23 or
Escherichia coli
ATCC 486 or against the producer strain itself.
N
-Dinitrophenyl derivatization and hydrazine hydrolysis of free amino and free carboxyl groups liberated from peptidoglycan digested with millericin B followed by thin-layer chromatography showed millericin B to be an endopeptidase with multiple activities. It cleaves the stem peptide at the N terminus of glutamic acid as well as the N terminus of the last residue in the interpeptide cross-link of susceptible strains.
Purification and Characterization of 2,6-β-
d
-Fructan 6-Levanbiohydrolase from
Streptomyces exfoliatus
F3-2
Authors: Saito Katsuichi , Kondo Kazuya , Kojima Ichiro , Yokota Atsushi , Tomita Fusao 
Published: April 2009

ABSTRACT

Streptomyces exfoliatus
F3-2 produced an extracellular enzyme that converted levan, a β-2,6-linked fructan, into levanbiose. The enzyme was purified 50-fold from culture supernatant to give a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weights of this enzyme were 54,000 by SDS-PAGE and 60,000 by gel filtration, suggesting the monomeric structure of the enzyme. The isoelectric point of the enzyme was determined to be 4.7. The optimal pH and temperature of the enzyme for levan degradation were pH 5.5 and 60°C, respectively. The enzyme was stable in the pH range 3.5 to 8.0 and also up to 50°C. The enzyme gave levanbiose as a major degradation product from levan in an exo-acting manner. It was also found that this enzyme catalyzed hydrolysis of such fructooligosaccharides as 1-kestose, nystose, and 1-fructosylnystose by liberating fructose. Thus, this enzyme appeared to hydrolyze not only β-2,6-linkage of levan, but also β-2,1-linkage of fructooligosaccharides. From these data, the enzyme from
S. exfoliatus
F3-2 was identified as a novel 2,6-β-
d
-fructan 6-levanbiohydrolase (EC
3.2.1.64
).
Detection and Characterization of Plasmid pJP4 Transfer to Indigenous Soil Bacteria
Authors: Newby D. T. , Josephson K. L. , Pepper I. L. 
Published: April 2009

ABSTRACT

Prior to gene transfer experiments performed with nonsterile soil, plasmid pJP4 was introduced into a donor microorganism,
Escherichia coli
ATCC 15224, by plate mating with
Ralstonia eutropha
JMP134. Genes on this plasmid encode mercury resistance and partial 2,4-dichlorophenoxyacetic acid (2,4-D) degradation. The
E. coli
donor lacks the chromosomal genes necessary for mineralization of 2,4-D, and this fact allows presumptive transconjugants obtained in gene transfer studies to be selected by plating on media containing 2,4-D as the carbon source. Use of this donor counterselection approach enabled detection of plasmid pJP4 transfer to indigenous populations in soils and under conditions where it had previously not been detected. In Madera Canyon soil, the sizes of the populations of presumptive indigenous transconjugants were 10
7
and 10
8
transconjugants g of dry soil
−1
for samples supplemented with 500 and 1,000 μg of 2,4-D g of dry soil
−1
, respectively. Enterobacterial repetitive intergenic consensus PCR analysis of transconjugants resulted in diverse molecular fingerprints. Biolog analysis showed that all of the transconjugants were members of the genus
Burkholderia
or the genus
Pseudomonas
. No mercury-resistant, 2,4-D-degrading microorganisms containing large plasmids or the
tfdB
gene were found in 2,4-D-amended uninoculated control microcosms. Thus, all of the 2,4-D-degrading isolates that contained a plasmid whose size was similar to the size of pJP4, contained the
tfdB
gene, and exhibited mercury resistance were considered transconjugants. In addition, slightly enhanced rates of 2,4-D degradation were observed at distinct times in soil that supported transconjugant populations compared to controls in which no gene transfer was detected.
Production and Comparison of Peptide Siderophores from Strains of Distantly Related Pathovars of
Pseudomonas syringae
and
Pseudomonas viridiflava
LMG 2352
Authors: Bultreys Alain , Gheysen Isabelle 
Published: April 2009

ABSTRACT

The production of peptide siderophores and the variation in siderophore production among strains of
Pseudomonas syringae
and
Pseudomonas viridiflava
were investigated. An antibiose test was used to select a free amino acid-containing agar medium favorable for production of fluorescent siderophores by two
P. syringae
strains. A culture technique in which both liquid and solid asparagine-containing culture media were used proved to be reproducible and highly effective for inducing production of siderophores in a liquid medium by the fluorescent
Pseudomonas
strains investigated. Using asparagine as a carbon source appeared to favor siderophore production, and relatively high levels of siderophores were produced when certain amino acids were used as the sole carbon and energy sources. Purified chelated siderophores of strains of
P. syringae
pv. syringae,
P. syringae
pv. aptata,
P. syringae
pv. morsprunorum,
P. syringae
pv. tomato, and
P. viridiflava
had the same amino acid composition and spectral characteristics and were indiscriminately used by these strains. In addition, nonfluorescent strains of
P. syringae
pv. aptata and
P. syringae
pv. morsprunorum were able to use the siderophores in biological tests. Our results confirmed the proximity of
P. syringae
and
P. viridiflava
; siderotyping between pathovars of
P. syringae
was not possible. We found that the spectral characteristics of the chelated peptide siderophores were different from the spectral characteristics of typical pyoverdins. Our results are discussed in relation to the ecology of the organisms and the conditions encountered on plant surfaces.
Endo-Xylogalacturonan Hydrolase, a Novel Pectinolytic Enzyme
Authors: van der Vlugt-Bergmans C. J. B. , Meeuwsen P. J. A. , Voragen A. G. J. , van Ooyen A. J. J. 
Published: April 2009

ABSTRACT

We screened an
Aspergillus tubingensis
expression library constructed in the yeast
Kluyveromyces lactis
for xylogalacturonan-hydrolyzing activity in microwell plates by using a bicinchoninic acid assay. This assay detects reducing carbohydrate groups when they are released from a carbohydrate by enzymatic activity. Two
K. lactis
recombinants exhibiting xylogalacturonan-hydrolyzing activity were found among the 3,400 colonies tested. The cDNA insert of these recombinants encoded a 406-amino-acid protein, designated XghA, which was encoded by a single-copy gene,
xghA
. A multiple-sequence alignment revealed that XghA was similar to both polygalacturonases (PGs) and rhamnogalacturonases. A detailed examination of conserved regions in the sequences of these enzymes revealed that XghA resembled PGs more. High-performance liquid chromatography and matrix-assisted laser desorption ionization–time of flight mass spectrometry of the products of degradation of xylogalacturonan and saponified modified hairy regions of apple pectin by XghA demonstrated that this enzyme uses an endo type of mechanism. XghA activity appeared to be specific for a xylose-substituted galacturonic acid backbone.