bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differentiation between
Candida dubliniensis
and
Candida albicans
by Fatty Acid Methyl Ester Analysis Using Gas-Liquid Chromatography
Authors: Peltroche-Llacsahuanga Heidrun , Schmidt Silke , Seibold Michael , Lütticken Rudolf , Haase Gerhard 
Published: December 2019

ABSTRACT

Candida dubliniensis
is often found in mixed culture with
C. albicans
, but its recognition is hampered as the color of its colonies in primary culture on CHROMagar Candida varies. Furthermore, definite identification of
C. dubliniensis
is difficult to achieve, time-consuming, and expensive. Therefore, a method to discriminate between these two closely related yeast species by fatty acid methyl ester (FAME) analysis using gas-liquid chromatography (Sherlock Microbial Identification System [MIS]; MIDI, Inc., Newark, Del.) was developed. Although the chromatograms of these two species revealed no obvious differences when applying FAME analysis, a new library (CADLIB) was successfully created using Sherlock Library Generation Software (MIDI). The amount and frequency of FAME was analyzed using library training files (
n
= 10 for each species), preferentially those comprising reference strains. For testing the performance of the CADLIB, clinical isolates genetically assigned to the respective species (
C. albicans
,
n
= 32;
C. dubliniensis
,
n
= 28) were chromatographically analyzed. For each isolate tested, MIS computed a similarity index (SI) indicating a hierarchy of possible strain fits. When using the newly created library CADLIB, the SIs for
C. albicans
and
C. dubliniensis
ranged from 0.11 to 0.96 and 0.53 to 0.93 (for all but one), respectively. Only three isolates of
C. albicans
(9.4%) were misidentified as
C. dubliniensis
, whereas all isolates of
C. dubliniensis
were correctly identified. Resulting differentiation accuracy was 90.6% for
C. albicans
and 100% for
C. dubliniensis
. Cluster analysis and principal component analysis of the resulting FAME profiles showed two clearly distinguishable clusters matching up with two assigned species for the strains tested. Thus, the created library proved to be well suited to discriminate between these two species.
Two Enzyme Immunoassays and PCR for Detection of
Helicobacter pylori
in Stool Specimens from Pediatric Patients before and after Eradication Therapy
Authors: Makristathis Athanasios , Barousch Wolfgang , Pasching Eva , Binder Christa , Kuderna Christa , Apfalter Petra , Rotter Manfred L. , Hirschl Alexander M. 
Published: December 2019

ABSTRACT

This study of pediatric patients was intended to determine the suitability of stool PCR and two antigen enzyme immunoassays (EIAs; Premier Platinum HpSA and the novel FemtoLab H. pylori), which detect
Helicobacter pylori
antigens in feces, as pretreatment diagnostic tools and especially as posttreatment control. Forty-nine
H. pylori
-infected children with dyspepsia received eradication therapy. Successful treatment was determined by a negative [
13
C]urea breath test 4 and 12 weeks after discontinuation of therapy. Fecal specimens were collected prior to eradication therapy as well as 4 weeks after the end of treatment. Successfully treated children delivered stool samples at 6, 8, and 12 weeks posttreatment also. Specimens were examined by seminested PCR and Premier Platinum HpSA and were reexamined by both EIAs as soon as FemtoLab H. pylori was available. In the first test series, the overall sensitivities of PCR and Premier Platinum HpSA were 93.0 and 91.1%, respectively. With specimens collected at 4 weeks after treatment, the respective specificities were 68.8 and 79.3%. After longer follow-up periods, however, they gradually increased to 100 and 96.9%, respectively. In the new test series, Premier Platinum HpSA delivered a considerably lower number of false-positive results (4 versus 18), indicating intertest variations. The overall test sensitivity was 94.6%, and the overall specificity was 97.5%. FemtoLab H. pylori showed an excellent performance with an overall sensitivity and specificity of 98.2 and 98.1%, respectively. Thus, in contrast to PCR, both EIAs were shown to be suitable for early posttreatment control.
Utilization of Exocellular Mannan from
Rhodotorula glutinis
as an Immunoreactive Antigen in Diagnosis of Leptospirosis
Authors: Matsuo Kouki , Isogai Emiko , Araki Yoshio 
Published: December 2019

ABSTRACT

Previously,
Rhodotorula glutinis
was reported to produce a large amount of exocellular mannan, having a repeating unit of →3)-
d
-Man
p
-(1→4)-
d
-Man
p
-(1→. Recently, we found that antigenic polysaccharides of
Leptospira biflexa
serovar patoc strain Patoc I have the same repeating unit and cross-react with antisera raised against extended strains of other leptospires (K. Matsuo, E. Isogai, and Y. Araki, Carbohydr. Res., in press). This structural identity and the difficulty of producing and isolating antigens led us to confirm the usefulness of
Rhodotorula
mannan as an immunoreactive antigen in a serological diagnosis of leptospirosis. In the present investigation, we confirmed the structural identity of an exocellular mannan isolated from
R. glutinis
AHU 3479 and tried to use it as an immunoreactive antigen in a serological diagnosis of leptospirosis. From its chemical analysis and
1
H- and
13
C-labeled nuclear magnetic resonance spectrometry, the
Rhodotorula
mannan was confirmed to consist of the same disaccharide units. Furthermore, such a preparation was shown to immunoreact to various sera from patients suffering with leptospirosis as well as to most rabbit antiserum preparations obtained from immunization with various strains of pathogenic leptospires. Therefore, the
Rhodotorula
mannan preparation is useful as an immunoreactive antigen in the serological diagnosis for leptospirosis.
Evaluation of a PCR Primer Based on the Isocitrate Dehydrogenase Gene for Detection of
Helicobacter pylori
in Feces
Authors: Argyros Frauke C. , Ghosh Mayurika , Huang Lili , Masubuchi Naoko , Cave David R. , Grübel Peter 
Published: December 2019

ABSTRACT

In order to improve detection and identification of
Helicobacter pylori
in highly contaminated samples, we evaluated new specific primers based on the DNA base sequence within the isocitrate dehydrogenase (
icd
) gene to amplify a 1,200-bp DNA segment. The specificity of the
icd
primer was tested against DNA derived from various bacteria, including 7
Helicobacter
species and a panel of 1 gram-variable, 2 gram-positive, and 16 gram-negative bacteria, as well as DNA from houseflies and feces from
H. pylori
-negative patients. The primers permitted the detection of all clinical
H. pylori
isolates tested, but no reactions were observed with negative controls. Several procedures for DNA extraction from feces were evaluated using PCR with
icd
primers. The lower limits of detection of
H. pylori
DNA from two different sources containing the same number of
H. pylori
organisms, a pure culture and feces spiked with
H. pylori
, were established for each extraction method tested. The results were 8.0 × 10
3
CFU/ml for cultures of pure
H. pylori
, and 8.0 × 10
6
CFU/ml for
H. pylori
from feces, using the phenol-chloroform method; 8.0 × 10
2
and 7.0 × 10
3
CFU/ml, respectively, for a glass matrix and chaotropic solution protocol; 8.0 × 10
2
and 7.0 × 10
3
CFU/ml, respectively, for the QIAamp tissue kit; and 5.0 × 10
2
and 5.0 × 10
3
CFU/ml, respectively, for the XTRAX DNA extraction kit. We conclude that the use of the
icd
gene as a primer for PCR represents a specific and sensitive assay for detection of
H. pylori
in highly contaminated samples.
Bacillus anthracis
Diversity in Kruger National Park
Authors: Smith K. L. , DeVos V. , Bryden H. , Price L. B. , Hugh-Jones M. E. , Keim P. 
Published: December 2019

ABSTRACT

The Kruger National Park (KNP), South Africa, has a recorded history of periodic anthrax epidemics causing widespread disease among wild animals.
Bacillus anthracis
is the causative agent of anthrax, a disease primarily affecting ungulate herbivores. Worldwide there is little diversity among
B. anthracis
isolates, but examination of variable-number tandem repeat (VNTR) loci has identified six major clones, with the most dissimilar types split into the A and B branches. Both the A and B types are found in southern Africa, giving this region the greatest genetic diversity of
B. anthracis
worldwide. Consequently, southern Africa has been hypothesized to be the geographic origin of
B. anthracis
. In this study, we identify the genotypic types of 98 KNP
B. anthracis
isolates using multiple-locus VNTR analysis. Two major types are evident, the A branch and the B branch. The spatial and temporal distribution of the different genotypes indicates that anthrax epidemic foci are independent, though correlated through environmental cues. Kruger B isolates were found on significantly higher-calcium and higher-pH soils than were Kruger type A. This relationship between genotype and soil chemistry may be due to adaptive differences among divergent anthrax strains. While this association may be simply fortuitous, adaptation of A types to diverse environmental conditions is consistent with their greater geographic dispersal and genetic dissimilarity.
Comparison between Plasma and Whole Blood Specimens for Detection of
Aspergillus
DNA by PCR
Authors: Loeffler Juergen , Hebart Holger , Brauchle Ulrike , Schumacher Ulrike , Einsele Hermann 
Published: December 2019

ABSTRACT

Ninety-six plasma and whole blood specimens from nine selected patients were analyzed for the presence of
Aspergillus
DNA. Nineteen specimens from three patients with proven aspergillosis were PCR positive in both materials, whereas an additional 22 were PCR positive in whole blood only. All 36 samples from six patients without signs of aspergillosis were negative in both assays. We conclude that although plasma and whole blood spiked with
Aspergillus
conidia showed an identical lower detection limit (10 CFU), the sensitivity of plasma PCR was lower than that of PCR performed on whole blood samples.
Isolation of
Moraxella canis
from an Ulcerated Metastatic Lymph Node
Authors: Vaneechoutte Mario , Claeys Geert , Steyaert Sophia , De Baere Thierry , Peleman Renaat , Verschraegen Gerda 
Published: December 2019

ABSTRACT

Moraxella canis
was isolated in large numbers from an ulcerated supraclavicular lymph node of a terminal patient, who died a few days later. Although the patient presented with septic symptoms and with a heavy growth of gram-negative diplococci in the lymph node, blood cultures remained negative.
M. canis
is an upper-airway commensal from dogs and cats and is considered nonpathogenic for humans, although this is the third reported human isolate of this species.
Mycobacterium branderi
from Both a Hand Infection and a Case of Pulmonary Disease
Authors: Wolfe Joyce , Turenne Christine , Alfa Michelle , Harding Godfrey , Thibert Louise , Kabani Amin 
Published: December 2019

ABSTRACT

Mycobacterium branderi
, a potential human pathogen first characterized in 1995, has been isolated from respiratory tract specimens. We report here a case in which
M. branderi
was the only organism isolated upon culture from a hand infection. This isolate, along with a second isolate from a bronchial specimen, was subjected to conventional identification tests for mycobacterial species. Further analysis by high-performance liquid chromatography (HPLC) of mycolic acids and 16S rRNA gene sequencing was performed, and the antibiotic susceptibility profile was determined for both strains. Biochemical tests and the HPLC pattern were consistent with that of
M. branderi
and
M. celatum
, which are very similar. The 16S rRNA gene sequence of both strains corresponded to that of
M. branderi
and enabled us to confidently differentiate this organism from other closely related species such as
M. celatum
. This contributes to a further understanding of the status of this species as a potential human pathogen as well as illustrating the need for molecular diagnostics as a complementary method for the identification of rare mycobacterial species.
Improved Immunodiagnosis of Cystic Hydatid Disease by Using a Synthetic Peptide with Higher Diagnostic Value Than That of Its Parent Protein,
Echinococcus granulosus
Antigen B
Authors: González-Sapienza Gualberto , Lorenzo Carmen , Nieto Alberto 
Published: December 2019

ABSTRACT
The assays are used for the diagnosis of hydatid disease are still imperfect. The reported diagnostic sensitivity and specificity vary greatly depending on the panel of sera used, the laboratory conducting the assay, and, more critically, the antigen used. To contribute to its standardization, we have recently ranked the diagnostic performances of the major parasite antigens and the available synthetic peptides using a large collection of serum samples. That work showed that antigen B (AgB) possesses the highest diagnostic value among these antigens. In the present work we further dissected its antigenicity by analyzing the reactivity of the same panel of sera against a set of synthetic peptides spanning the sequence of both AgB subunits. The N-terminal extension of these subunits appeared to be immunodominant in human infections. A 38-mer peptide (p176) delineated from the N-terminal extension of the AgB/1 subunit performed in an enzyme-linked immunosorbent assay with a higher diagnostic sensitivity (80%) and specificity (94%) than native AgB, Ag5, or any other peptide antigen tested against this collection of serum samples. In view of its high diagnostic value and its nature as a well-defined reproducible antigen, p176 could conveniently be used as a reference standard antigen in the diagnosis of hydatid disease.
Evaluation of Fluorescence-Based Amplified Fragment Length Polymorphism Analysis for Molecular Typing in Hospital Epidemiology: Comparison with Pulsed-Field Gel Electrophoresis for Typing Strains of Vancomycin-Resistant
Enterococcus faecium
Authors: Antonishyn Nick A. , McDonald Ryan R. , Chan Edward L. , Horsman Greg , Woodmansee Carla E. , Falk Pamela S. , Mayhall C. Glen 
Published: December 2019

ABSTRACT
Fluorescence-based amplified fragment length polymorphism (fbAFLP) is a novel assay based on the fluorescent analysis of an amplified subset of restriction fragments. The fbAFLP assay involves the selective PCR amplification of restriction fragments from a total digest of genomic DNA. The ligation of adapters with primer-specific sites coupled with primers containing selective nucleotides allowed the full potential of PCR to be realized while maintaining the advantages of restriction endonuclease analysis. Fluorescence-based fragment analysis with polyacrylamide gel electrophoresis provides the accurate band sizing required for homology assessment. The large number of phylogenetically informative characters obtained by fbAFLP is well suited for cluster analysis and database development. The method demonstrated excellent reproducibility and ease of performance and interpretation. We typed 30 epidemiologically well-characterized isolates of vancomycin-resistant enterococci from an outbreak in a university hospital by fbAFLP. Clustering of fbAFLP data matched epidemiological, microbiological, and pulsed-field gel electrophoresis data. This study demonstrates the unprecedented utility of fbAFLP for epidemiological investigation. Future developments in standardization and automation will set fbAFLP as the “gold standard” for molecular typing in epidemiology.
Detection and Identification of Mycobacteria by Amplification of the Internal Transcribed Spacer Regions with Genus- and Species-Specific PCR Primers
Authors: Park Heekyung , Jang Hyunjung , Kim Cheolmin , Chung Byungseon , Chang Chulhun L. , Park Soon Kew , Song Sundae 
Published: December 2019

ABSTRACT

We evaluated the usefulness of PCR assays that target the internal transcribed spacer (ITS) region for identifying mycobacteria at the species level. The conservative and species-specific ITS sequences of 33 species of mycobacteria were analyzed in a multialignment analysis. One pair of panmycobacterial primers and seven pairs of mycobacterial species-specific primers were designed. All PCRs were performed under the same conditions. The specificities of the primers were tested with type strains of 20 mycobacterial species from the American Type Culture Collection; 205 clinical isolates of mycobacteria, including 118
Mycobacterium tuberculosis
isolates and 87 isolates of nontuberculous mycobacteria from 10 species; and 76 clinical isolates of 28 nonmycobacterial pathogenic bacterial species. PCR with the panmycobacterial primers amplified fragments of approximately 270 to 400 bp in all mycobacteria. PCR with the
M. tuberculosis
complex-specific primers amplified an approximately 120-bp fragment only for the
M. tuberculosis
complex. Multiplex PCR with the panmycobacterial primers and the
M. tuberculosis
complex-specific primers amplified two fragments that were specific for all mycobacteria and the
M. tuberculosis
complex, respectively. PCR with
M. avium
complex-,
M. fortuitum-, M. chelonae-, M. gordonae-, M. scrofulaceum-
, and
M. szulgai
-specific primers amplified specific fragments only for the respective target organisms. These novel primers can be used to detect and identify mycobacteria simultaneously under the same PCR conditions. Furthermore, this protocol facilitates early and accurate diagnosis of mycobacteriosis.
Mycobacterium heckeshornense
sp. nov., a New Pathogenic Slowly Growing
Mycobacterium
sp. Causing Cavitary Lung Disease in an Immunocompetent Patient
Authors: Roth Andreas , Reischl Udo , Schönfeld Nicolas , Naumann Ludmila , Emler Stefan , Fischer Marga , Mauch Harald , Loddenkemper Robert , Kroppenstedt Reiner M. 
Published: December 2019

ABSTRACT

A pathogenic scotochromogenic
Mycobacterium xenopi
-like organism was isolated from the lung of an immunocompetent young woman. This pathogen caused severe bilateral cavitary lung disease, making two surgical interventions necessary after years of chronic disease. This case prompted us to characterize this mycobacterium by a polyphasic taxonomic approach. The isolate contained chemotaxonomic markers which were typical for the genus
Mycobacterium
, i.e., the
meso
isomer of 2,6-diaminopimelic acid, arabinose, and galactose as diagnostic whole-cell sugars, MK-9(H
2
) as the principal isoprenoid quinone, a mycolic acid pattern of α-mycolates, ketomycolates, and wax ester mycolates, unbranched saturated and unsaturated fatty acids plus a significant amount of tuberculostearic acid, and small amounts of a C
20:0
secondary alcohol. On the basis of its unique 16S rRNA and 16S-23S spacer gene sequences, we propose that the isolate should be assigned to a new species,
Mycobacterium heckeshornense
. This novel species is phylogenetically closely related to
M. xenopi
. The type strain of
M. heckeshornense
is strain S369 (DSM 44428
T
). The GenBank accession number of the 16S rRNA gene of
M. heckeshornense
is
AF174290
.
PCR Detection of
Escherichia coli
O157:H7 Directly from Stools: Evaluation of Commercial Extraction Methods for Purifying Fecal DNA
Authors: Holland J. L. , Louie L. , Simor A. E. , Louie M. 
Published: December 2019

ABSTRACT

Rapid identification of
Escherichia coli
O157:H7 is important for patient management and for prompt epidemiological investigations. We evaluated one in-house method and three commercially available kits for their ability to extract
E. coli
O157:H7 DNA directly from stool specimens for PCR. Of the 153 stool specimens tested, 107 were culture positive and 46 were culture negative. The sensitivities and specificities of the in-house enrichment method, IsoQuick kit, NucliSens kit, and QIAamp kit were comparable, as follows: 83 and 98%, 85 and 100%, 74 and 98%, and 86 and 100%, respectively. False-negative PCR results may be due to the presence of either inherent inhibitors or small numbers of organisms. The presence of large amounts of bacteria relative to the amount of the
E. coli
O157:H7 target may result in the lower sensitivities of the assays. All commercial kits were rapid and easy to use, although DNA extracted with the QIAamp kit did not require further dilution of the DNA template prior to PCR.
Improved Version 2.0 Qualitative and Quantitative AMPLICOR Reverse Transcription-PCR Tests for Hepatitis C Virus RNA: Calibration to International Units, Enhanced Genotype Reactivity, and Performance Characteristics
Authors: Lee Sung C. , Antony Anisha , Lee Nitta , Leibow Jeff , Yang Jian Q. , Soviero Stephen , Gutekunst Karen , Rosenstraus Maurice 
Published: December 2019

ABSTRACT
Version 2.0 qualitative and quantitative AMPLICOR reverse transcription-PCR tests for HCV were designed to improve on the performance of first version of the hepatitis C virus (HCV) tests. The new tests were calibrated in international units, the new commonly accepted standard unit of measurement for HCV RNA. The sensitivity of the qualitative tests was enhanced by modifying the specimen processing procedure to achieve a limit of detection 50 IU/ml. The limit of detection for the quantitative tests was 600 IU/ml. Modifications to the amplification reaction mixture and thermal cycling conditions enabled all genotypes to be amplified with similar efficiency. The quantitative tests exhibited a linear range extending from 500 to 500,000 IU/ml and excellent reproducibility, with coefficients of variation ranging from 18 to 39%, within the linear range. These data indicate that the version 2.0 AMPLICOR HCV tests will improve diagnosis of HCV infection and will yield more-accurate titers for prognosis and for monitoring therapeutic efficacy, particularly at low viral loads. Furthermore, it will be possible to compare the performance characteristics and viral load measurements of AMPLICOR tests to those of other tests that adopt the international unit as the standard of measurement.
Panfungal PCR and Multiplex Liquid Hybridization for Detection of Fungi in Tissue Specimens
Authors: Hendolin Panu H. , Paulin Lars , Koukila-Kähkölä Pirkko , Anttila Veli-Jukka , Malmberg Henrik , Richardson Malcolm , Ylikoski Jukka 
Published: December 2019

ABSTRACT

A procedure based on panfungal PCR and multiplex liquid hybridization was developed for the detection of fungi in tissue specimens. The PCR amplified the fungal internal transcribed spacer (ITS) region (ITS1-5.8S rRNA-ITS2). After capture with specific probes, eight common fungal pathogens (
Aspergillus flavus
,
Aspergillus fumigatus
,
Candida albicans
,
Candida krusei
,
Candida glabrata
,
Candida parapsilosis
,
Candida tropicalis
, and
Cryptococcus neoformans
) were identified according to the size of the amplification product on an automated sequencer. The nonhybridized products were identified by sequencing. The performance of the procedure was examined with 12 deep-tissue specimens and 8 polypous tissue biopsies from the paranasal sinuses. A detection level of 0.1 to 1 pg of purified DNA (2 to 20 CFU) was achieved. Of the 20 specimens, PCR was positive for 19 (95%), of which 10 (53%) were hybridization positive. In comparison, 12 (60%) of the specimens were positive by direct microscopy, but only 7 (35%) of the specimens showed fungal growth. Sequencing of the nonhybridized amplification products identified an infecting agent in six specimens, and three specimens yielded only sequences of unknown fungal origin. The procedure provides a rapid (within 2 days) detection of common fungal pathogens in tissue specimens, and it is highly versatile for the identification of other fungal pathogens.