bluuebunny/crossref_metadata_2025_split huggingface dataset:
Rapid and Simple Determination of Ciprofloxacin Resistance in Clinical Strains of
Escherichia coli
Authors: Santiso Rebeca , Tamayo María , Fernández José Luis , del Carmen Fernández María , Molina Francisca , Villanueva Rosa , Gosálvez Jaime , Bou Germán 
Published: July 2009

ABSTRACT

We recently reported a simple new in situ diffusion assay, developed as a kit, to visualize DNA fragmentation in single bacterial cells. Use of this assay in a collection of 95 genetically unrelated
Escherichia coli
clinical strains resulted in correct identification of all of the isolates as resistant or susceptible to ciprofloxacin, consistent with the MIC results. This relevant information is obtained in 80 min.
Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Fast and Reliable Identification of Clinical Yeast Isolates
Authors: Marklein G. , Josten M. , Klanke U. , Müller E. , Horré R. , Maier T. , Wenzel T. , Kostrzewa M. , Bierbaum G. , Hoerauf A. , Sahl H.-G. 
Published: July 2009

ABSTRACT

The clinical impact of severe infections with yeasts and yeast-like fungi has increased, especially in immunocompromised hosts. In recent years, new antifungal agents with different and partially species-specific activity patterns have become available. Therefore, rapid and reliable species identification is essential for antifungal treatment; however, conventional biochemical methods are time-consuming and require considerable expertise. We evaluated matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for the rapid routine identification of clinical yeast isolates. A total of 18 type collection strains and 267 recent clinical isolates of
Candida
(
n
= 250),
Cryptococcus
,
Saccharomyces
,
Trichosporon
,
Geotrichum
,
Pichia
, and
Blastoschizomyces
spp. were identified by MALDI-TOF MS. The results were compared with those obtained by conventional phenotyping and biochemical tests, including the API ID 32C system (bioMérieux, Nürtingen, Germany). Starting with cells from single colonies, accurate species identification by MALDI-TOF MS was achieved for 247 of the clinical isolates (92.5%). The remaining 20 isolates required complementation of the reference database with spectra for the appropriate reference strains which were obtained from type culture collections or identified by 26S rRNA gene sequencing. The absence of a suitable reference strain from the MALDI-TOF MS database was clearly indicated by log(score) values too low for the respective clinical isolates; i.e., no false-positive identifications occurred. After complementation of the database, all isolates were unambiguously identified. The established API ID 32C biochemical diagnostic system identified 244 isolates in the first round. Overall, MALDI-TOF MS proved a most rapid and reliable tool for the identification of yeasts and yeast-like fungi, with the method providing a combination of the lowest expenditure of consumables, easy interpretation of results, and a fast turnaround time.
Evaluation of Quinolones for Use in Detection of Determinants of Acquired Quinolone Resistance, Including the New Transmissible Resistance Mechanisms
qnrA
,
qnrB
,
qnrS
, and
aac
(6′)
Ib-cr
, in
Escherichia coli
and
Salmonella enterica
and Determinations of Wild-Type Distributions
Authors: Cavaco L. M. , Aarestrup F. M. 
Published: July 2009

ABSTRACT

Fluoroquinolone resistance in members of the
Enterobacteriaceae
family is mostly due to mutations in the quinolone resistance-determining regions of the topoisomerase genes. However, transferable genes encoding quinolone resistance have recently been described. The current methods for susceptibility testing are not adapted to the detection of new resistance determinants, which confer low levels of resistance. The aim of this study was to compare the ability of the screening of the different quinolones by disk diffusion assays and MIC determinations to detect fluoroquinolone resistance. Sixty-nine
Escherichia coli
strains and 62
Salmonella
strains, including strains fully susceptible to quinolones, nalidixic acid-resistant strains, strains with resistance to fluoroquinolones (resistant to nalidixic acid), and strains showing low-level resistance to fluoroquinolones conferred by transferable quinolone resistance genes, including
qnrA
,
qnrB
,
qnrS
, and
aac
(6′)
Ib-cr
, were selected. Disk diffusion assays and MIC determinations by the agar dilution method were performed, according to CLSI standards, with nalidixic acid, flumequine, oxolinic acid, ciprofloxacin, enrofloxacin, marbofloxacin, norfloxacin, ofloxacin, and levofloxacin. The MIC of levofloxacin was determined by an Etest. The results showed a trimodal distribution of the MICs for both
E. coli
and
Salmonella
. The MIC distributions for the isolates varied with the compounds tested. Screening for nalidixic acid resistance by MIC testing or disk diffusion assay was not efficient for the detection of some of the isolates carrying
qnr
and
aac
(6′)
Ib-cr
. Transferable resistance genes would best be detected by testing for the MIC of ciprofloxacin or norfloxacin, as testing for the MICs of the other compounds would fail to detect isolates carrying
aac
(6′)
Ib-cr
because the enzyme produced is able to reduce the activities of these two compounds only due to their chemical structures. In conclusion, screening with nalidixic acid is efficient for the detection of mutants, but it is not so efficient for the detection of
qnr
and
aac
(6′)
Ib-cr
. Detection would be maximized by screening with either ciprofloxacin or norfloxacin by both MIC determination and disk diffusion assays. Furthermore, a low concentration of ciprofloxacin (1 μg) in the disks seemed to increase the sensitivity of the disk diffusion assay.
Utility of a Commercially Available Multiplex Real-Time PCR Assay To Detect Bacterial and Fungal Pathogens in Febrile Neutropenia
Authors: von Lilienfeld-Toal Marie , Lehmann Lutz E. , Raadts Ansgar D. , Hahn-Ast Corinna , Orlopp Katjana S. , Marklein Günter , Purr Ingvill , Cook Gordon , Hoeft Andreas , Glasmacher Axel , Stüber Frank 
Published: July 2009

ABSTRACT

Infection is the main treatment-related cause of mortality in cancer patients. Rapid and accurate diagnosis to facilitate specific therapy of febrile neutropenia is therefore urgently warranted. Here, we evaluated a commercial PCR-based kit to detect the DNA of 20 different pathogens (SeptiFast) in the setting of febrile neutropenia after chemotherapy. Seven hundred eighty-four serum samples of 119 febrile neutropenic episodes (FNEs) in 70 patients with hematological malignancies were analyzed and compared with clinical, microbiological, and biochemical findings. In the antibiotic-naïve setting, bacteremia was diagnosed in 34 FNEs and 11 of them yielded the same result in the PCR. Seventy-three FNEs were negative in both systems, leading to an overall agreement in 84 of 119 FNEs (71%). During antibiotic therapy, positivity in blood culture occurred only in 3% of cases, but the PCR yielded a positive result in 15% of cases. In six cases the PCR during antibiotic treatment detected a new pathogen repetitively; this was accompanied by a significant rise in procalcitonin levels, suggestive of a true detection of infection. All patients with probable invasive fungal infection (IFI;
n
= 3) according to the standards of the European Organization for Research and Treatment of Cancer had a positive PCR result for
Aspergillus fumigatus
; in contrast there was only one positive result for
Aspergillus fumigatus
in an episode without signs and symptoms of IFI. Our results demonstrate that the SeptiFast kit cannot replace blood cultures in the diagnostic workup of FNEs. However, it might be helpful in situations where blood cultures remain negative (e.g., during antimicrobial therapy or in IFI).
Emergence of a Community-Associated Methicillin-Resistant
Staphylococcus aureus
Strain with a Unique Resistance Profile in Southwest Nigeria
Authors: Ghebremedhin B. , Olugbosi M. O. , Raji A. M. , Layer F. , Bakare R. A. , König B. , König W. 
Published: July 2009

ABSTRACT

Phenotypic, genotypic, and toxin gene analyses have not yet been done all in one for the Nigerian
Staphylococcus aureus
population. This study provides a comprehensive overview of the molecular epidemiology and genetic diversity of
S. aureus
strains at the largest university clinic in Ibadan, Nigeria. From 1,300 patients' clinical samples collected at the University Teaching Hospital in Ibadan, Nigeria, during a 1-year-surveillance in 2007, 346 nonduplicate
S. aureus
isolates were obtained. All isolates underwent antibiotic susceptibility testing, toxin gene analysis, multilocus sequence typing,
agr
group typing, and
spa
typing. For methicillin (meticillin)-resistant
S. aureus
(MRSA), staphylococcal cassette chromosome
mec
(SCC
mec
) typing was also performed. Of the 346 isolates, 20.23% were methicillin resistant. Thirty-three patients' isolates (47.15%) fulfilled the definition criteria for community-associated MRSA (CA-MRSA) according to a review of the medical charts. The majority of MRSA strains analyzed were isolated from surgical or pediatric patients. The commonest types of MRSA infection identified were surgical-site infections (>70%), whereas those for CA-MRSA were conjunctivitis and otitis (19 patients [57.6%]) and accidental skin and subcutaneous tissue infections (14 patients [42.4%]). The methicillin-susceptible
S. aureus
strains (ST1, ST5, ST15, ST7, ST8, ST25, ST30, ST72, ST80, ST121, and ST508) were heterogeneous by phenotypic and genotypic analyses. The first report of a Panton-Valentine leukocidin-positive ST88 strain (
agr
III, SCC
mec
IV) in Nigeria, as well as genetic analyses of this strain, is presented in this study. The ST88 strain was resistant to trimethoprim-sulfamethoxazole as well as to penicillin and oxacillin. CA-MRSA infections are increasing rapidly among young patients with ophthalmologic and auricular infections. Urban regions with populations of lower socioeconomic status and evidence of overcrowding appear to be at high risk for the emergence of this clone.
A Trilocus Sequence Typing Scheme for Hospital Epidemiology and Subspecies Differentiation of an Important Nosocomial Pathogen,
Enterococcus faecalis
Authors: Chowdhury Shahreen A. , Arias Cesar A. , Nallapareddy Sreedhar R. , Reyes Jinnethe , Willems Rob J. L. , Murray Barbara E. 
Published: July 2009

ABSTRACT

In this study, we present a trilocus sequence typing (TLST) scheme based on intragenic regions of two antigenic genes,
ace
and
salA
(encoding a collagen/laminin adhesin and a cell wall-associated antigen, respectively), and a gene associated with antibiotic resistance,
lsa
(encoding a putative ABC transporter), for subspecies differentiation of
Enterococcus faecalis
. Each of the alleles was analyzed using 50
E. faecalis
isolates representing 42 diverse multilocus sequence types (ST
M
; based on seven housekeeping genes) and four groups of clonally linked (by pulsed-field gel electrophoresis [PFGE]) isolates. The allelic profiles and/or concatenated sequences of the three genes agreed with multilocus sequence typing (MLST) results for typing of 49 of the 50 isolates; in addition to the one exception, two isolates were found to have identical TLST types but were single-locus variants (differing by a single nucleotide) by MLST and were therefore also classified as clonally related by MLST. TLST was also comparable to PFGE for establishing short-term epidemiological relationships, typing all isolates classified as clonally related by PFGE with the same type. TLST was then applied to representative isolates (of each PFGE subtype and isolation year) of a collection of 48 hospital isolates and demonstrated the same relationships between isolates of an outbreak strain as those found by MLST and PFGE. In conclusion, the TLST scheme described here was shown to be successful for investigating short-term epidemiology in a hospital setting and may provide an alternative to MLST for discriminating isolates.
Two Cases of
Mycobacterium microti
-Induced Culture-Negative Tuberculosis
Authors: de Jong Eefje , Rentenaar Rob J. , van Pelt Rob , de Lange Wiel , Schreurs Wim , van Soolingen Dick , Sturm Patrick D. J. 
Published: July 2009

ABSTRACT

We present descriptions of two human
Mycobacterium microti
infections: one of a patient with pulmonary disease and one of a patient with osteomyelitis of the hip. Both patients had acid-fast bacilli and a positive
Mycobacterium tuberculosis
complex PCR from clinical specimens, but mycobacterial cultures remained negative. The microbiological diagnosis was established by molecular methods.
Characterization of Mycobacteria from a Major Brazilian Outbreak Suggests that Revision of the Taxonomic Status of Members of the
Mycobacterium chelonae
-
M. abscessus
Group Is Needed
Authors: Leao Sylvia Cardoso , Tortoli Enrico , Viana-Niero Cristina , Ueki Suely Yoko Mizuka , Lima Karla Valeria Batista , Lopes Maria Luiza , Yubero Jesus , Menendez Maria Carmen , Garcia Maria Jesus 
Published: July 2009

ABSTRACT

An outbreak of postsurgical infections caused by rapidly growing mycobacteria has been ongoing in Brazil since 2004. The degrees of similarity of the
rpoB
and
hsp65
sequences from the clinical isolates and the corresponding sequences from both the
Mycobacterium massiliense
and the
M. bolletii
type strains were above the accepted limit for interspecies variability, leading to conflicting identification results. Therefore, an extensive characterization of members of the
M. chelonae-M. abscessus
group was carried out. The
M. abscessus
,
M. chelonae
,
M. immunogenum
,
M. massiliense
, and
M. bolletii
type strains and a subset of clinical isolates were analyzed by biochemical tests, high-performance liquid chromatography, drug susceptibility testing, PCR-restriction enzyme analysis of
hsp65
(PRA-
hsp65
),
rpoB
, and
hsp65
gene sequencing and analysis of phylogenetic trees, DNA-DNA hybridization (DDH), and restriction fragment length polymorphism (RFLP) analysis of the 16S rRNA gene (RFLP-16S rRNA). The clinical isolates and the
M. abscessus
,
M. massiliense
, and
M. bolletii
type strains could not be separated by phenotypic tests and were grouped in the phylogenetic trees obtained. The results of DDH also confirmed the >70% relatedness of the clinical isolates and the
M. abscessus
,
M. massiliense
, and
M. bolletii
type strains; and indistinguishable RFLP-16S rRNA patterns were obtained. On the contrary, the separation of clinical isolates and the
M. abscessus
,
M. massiliense
, and
M. bolletii
type strains from
M. chelonae
and
M. immunogenum
was supported by the results of PRA-
hsp65
, DDH, and RFLP-16S rRNA and by the
rpoB
and
hsp65
phylogenetic trees. Taken together, these results led to the proposition that
M. abscessus
,
M. massiliense
, and
M. bolletii
represent a single species, that of
M. abscessus.
Two subspecies are also proposed,
M. abscessus
subsp.
abscessus
and
M. abscessus
subsp.
massiliense
, and these two subspecies can be distinguished by two different PRA-
hsp65
patterns, which differ by a single HaeIII band, and by differences in their
rpoB
(3.4%) and
hsp
65 (1.3%) sequences.
Identification of 20 Common Human Enterovirus Serotypes by Use of a Reverse Transcription-PCR-Based Reverse Line Blot Hybridization Assay
Authors: Zhou Fei , Kong Fanrong , McPhie Kenneth , Ratnamohan Mala , Donovan Linda , Zeng Frank , Gilbert Gwendolyn L. , Dwyer Dominic E. 
Published: July 2009

ABSTRACT
The more than 100 human enterovirus (HEV) serotypes can also be classified into four species, HEV-A to -D, based on phylogenetic analysis of multiple gene regions. Current molecular typing methods depend largely on reverse transcription-PCR (RT-PCR) amplification and nucleotide sequencing of the entire or 3′ half of the VP1 gene. An RT-PCR-based reverse line blot (RLB) hybridization assay was developed as a rapid and efficient approach to characterize common HEVs. Twenty HEV serotypes accounted for 87.1% of all HEVs isolated at an Australian reference virology laboratory from 1979 to 2007. VP1 sequences of all known HEV prototype strains were aligned to design one sense primer and three antisense primers for RT-PCR. After sequencing of the complete VP1 genes of 37 previously serotyped examples of the commonest 20 serotypes and alignment of these VP1 sequences with GenBank sequences, four serotype-specific probes for each serotype were designed for RLB. The RT-PCR-RLB assay was then applied to 132 HEV isolates, made up of the previously sequenced 37 isolates and another 95 serotyped clinical isolates. The RT-PCR-RLB genotypes corresponded with the serotypes for 131/132 isolates; the one exception was confirmed by VP1 sequencing, and the genotype was confirmed by repeat conventional serotyping. Genotyping by RT-PCR-RLB complements traditional serotyping methods and VP1 sequencing and has the advantages of convenience, speed, and accuracy. RT-PCR-RLB allows detection of specific enteroviral serotypes or genotypes associated with HEV outbreaks and significant disease.
WHO Global Salm-Surv External Quality Assurance System for Serotyping of
Salmonella
Isolates from 2000 to 2007
Authors: Hendriksen Rene S. , Mikoleit Matthew , Carlson Valeria P. , Karlsmose Susanne , Vieira Antonio R. , Jensen Arne B. , Seyfarth Anne Mette , DeLong Stephanie M. , Weill François-Xavier , Lo Fo Wong Danilo Marino Armando , Angulo Frederick J. , Wegener Henrik C. , Aarestrup Frank M. 
Published: July 2009

ABSTRACT

An international external quality assurance system (EQAS) for the serotyping of
Salmonella
species was initiated in 2000 by WHO Global Salm-Surv to enhance the capacity of national reference laboratories to obtain reliable data for surveillance purposes worldwide. Seven EQAS iterations were conducted between 2000 and 2007. In each iteration, participating laboratories submitted serotyping results for eight
Salmonella
isolates. A total of 249 laboratories in 96 countries participated in at least one EQAS iteration. A total of 756 reports were received from the participating laboratories during the seven EQAS iterations. Cumulatively, 76% of participating laboratories submitted data for all eight strains, and 82% of strains were correctly serotyped. In each iteration, 84% to 96% of the laboratories correctly serotyped the
Salmonella enterica
serovar Enteritidis isolate that was included as an internal quality control strain. Regional differences in performance were observed, with laboratories in Central Asia and the Middle East performing less well overall than those in other regions. Errors that resulted in incorrect serovar identification were typically caused by difficulties in the detection of the phase two flagellar antigen or in differentiation within antigen complexes; some of these errors are likely related to the quality of the antisera available. The results from the WHO Global Salm-Surv EQAS, the largest of its kind in the world, show that most laboratories worldwide are capable of correctly serotyping
Salmonella
species. However, this study also indicates a continuing need for improvement. Future training efforts should be aimed at enhancing the ability to detect the phase two flagellar antigen and at disseminating information on where to purchase high-quality antisera.
Simian Virus 40 T-Antigen-Mediated Gene Regulation in Enterocytes Is Controlled Primarily by the Rb-E2F Pathway
Authors: Rathi Abhilasha V. , Sáenz Robles M. Teresa , Cantalupo Paul G. , Whitehead Robert H. , Pipas James M. 
Published: July 2009

ABSTRACT

Simian virus 40 large T antigen (TAg) contributes to cell transformation, in part, by targeting two well-characterized tumor suppressors, pRb and p53. TAg expression affects the transcriptional circuits controlled by Rb and by p53. We have performed a microarray analysis to examine the global change in gene expression induced by wild-type TAg (TAg
wt
) and TAg mutants, in an effort to link changes in gene expression to specific transforming functions. For this analysis we have used enterocytes from the mouse small intestine expressing TAg. Expression of TAg
wt
in the mouse intestine results in hyperplasia and dysplasia. Our analysis indicates that practically all gene expression regulated by TAg in enterocytes is dependent upon its binding and inactivation of the Rb family proteins. To further dissect the role of the Rb family in the induction of intestinal hyperplasia, we have screened several lines of transgenic mice expressing a truncated TAg (TAg
N136
), which is able to interfere with the Rb pathway but lacks the functions associated with the carboxy terminus of the protein. This analysis confirmed the pivotal association between the Rb pathway and the induction of intestinal hyperplasia and revealed that upregulation of p53 target genes is not associated with the tumorigenic phenotype. Furthermore, we found that TAg
N136
was sufficient to induce intestinal hyperplasia, although the appearance of dysplasia was significantly delayed.
Human Immunodeficiency Virus Type 1 Escape from Cyclotriazadisulfonamide-Induced CD4-Targeted Entry Inhibition Is Associated with Increased Neutralizing Antibody Susceptibility
Authors: Vermeire Kurt , Van Laethem Kristel , Janssens Wouter , Bell Thomas W. , Schols Dominique 
Published: July 2009

ABSTRACT
Continuous specific downmodulation of CD4 receptor expression in T lymphocytes by the small molecule cyclotriazadisulfonamide (CADA) selected for the CADA-resistant human immunodeficiency virus type 1 (HIV-1) NL4.3 virus containing unique mutations in the C4 and V5 regions of gp120, likely stabilizing the CD4-binding conformation. The amino acid changes in Env were associated with decreased susceptibility to anti-CD4 monoclonal antibody treatment of the cells and with higher susceptibility of the virus to soluble CD4. In addition, the acquired ability of a CADA-resistant virus to infect cells with low CD4 expression was associated with an increased susceptibility of the virus to neutralizing antibodies from sera of several HIV-1-infected patients.
The Flexible Loop of the Human Cytomegalovirus DNA Polymerase Processivity Factor ppUL44 Is Required for Efficient DNA Binding and Replication in Cells
Authors: Alvisi Gualtiero , Roth Daniela Martino , Camozzi Daria , Pari Gregory S. , Loregian Arianna , Ripalti Alessandro , Jans David A. 
Published: July 2009

ABSTRACT

Phosphoprotein ppUL44 of the human cytomegalovirus (HCMV) DNA polymerase plays an essential role in viral replication, conferring processivity to the DNA polymerase catalytic subunit pUL54 by tethering it to the DNA. Here, for the first time, we examine in living cells the function of the highly flexible loop of ppUL44 (UL44-FL; residues 162 to 174 [PHTRVKRNVKKAP
174
]), which has been proposed to be directly involved in ppUL44's interaction with DNA. In particular, we use a variety of approaches in transfected cells to characterize in detail the behavior of ppUL44Δloop, a mutant derivative in which three of the five basic residues within UL44-FL are replaced by nonbasic amino acids. Our results indicate that ppUL44Δloop is functional in dimerization and binding to pUL54 but strongly impaired in binding nuclear structures within the nucleus, as shown by its inability to form nuclear speckles, reduced nuclear accumulation, and increased intranuclear mobility compared to wild-type ppUL44. Moreover, analysis of cellular fractions after detergent and DNase treatment indicates that ppUL44Δloop is strongly reduced in DNA-binding ability, in similar fashion to ppUL44-L86A/L87A, a point mutant derivative impaired in dimerization. Finally, ppUL44Δloop fails to transcomplement HCMV
ori
Lyt-dependent DNA replication in cells and also inhibits replication in the presence of wild-type ppUL44, possibly via formation of heterodimers defective for double-stranded DNA binding. UL44-FL thus emerges for the first time as an important determinant for HCMV replication in cells, with potential implications for the development of novel antiviral approaches by targeting HCMV replication.