bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Efficacy of Laboratory Diagnosis of
Helicobacter pylori
Infections in Gastric Biopsy Specimens Is Related to Bacterial Density and
vacA
,
cagA
, and
iceA
Genotypes
Authors: van Doorn Leen-Jan , Henskens Yvonne , Nouhan Nathalie , Verschuuren Anita , Vreede Rolf , Herbink Paul , Ponjee Gabriëlle , van Krimpen Kees , Blankenburg Ruud , Scherpenisse Joost , Quint Wim 
Published: May 2021

ABSTRACT

A total of 500 consecutive patients undergoing upper endoscopy were biopsied and tested for
H. pylori
infection by the
Campylobacter
-like organism (CLO) test, culture, histology, and PCR. Serum samples were tested by two different serological assays. Patients were considered
H. pylori
positive if at least two of the four biopsy specimen-based methods yielded positive results. PCR had the highest diagnostic sensitivity (99.4%), followed by histology (92.2%), culture (89.5%), and the CLO test (89.0%). The specificities of all methods were higher than 98%. Of the organisms from the 181 PCR-positive patients, the
vacA
(s and m regions),
cagA
, and
iceA
genotypes were determined by reverse hybridization (line probe assay) or an allele-specific PCR. Organisms that were detected by PCR but that remained undetected by the CLO test were significantly more often
vacA
s1 (
P
= 0.006), m1 (
P
= 0.028), and
cagA
positive (
P
= 0.029) than
vacA
s2, m2, and
cagA
negative, respectively. Organisms that were detected by PCR but that remained undetected by culture or histology more often contained
iceA1
(
P
= 0.034 and
P
= 0.029, respectively) than
iceA2
. Higher
H. pylori
density was associated with
vacA
s2 (
P
= 0.024),
vacA
m2 (
P
= 0.050), and
cagA
-negative (
P
= 0.035) genotypes. Also, the diagnostic results of the CLO test (
P
= 0.001) and culture (
P
= 0.031) but not those of the PCR (
P
= 0.130) were significantly associated with the
H. pylori
density. The rate of detection by the four biopsy specimen-based tests was lower for patients who used proton pump inhibitors, but this was independent of the
H. pylori
genotypes. These observations may be explained by different bacterial densities, as established by the distinct genotypes of
H. pylori
, and confirm that the biologies of strains with such genotypes are considerably different.
Specific
Taenia crassiceps
and
Taenia solium
Antigenic Peptides for Neurocysticercosis Immunodiagnosis Using Serum Samples
Authors: Bueno Ednéia Casagranda , Vaz Adelaide José , Machado Luís Dos Ramos , Livramento JoséAntônio , Mielle Sílvia Regina 
Published: May 2021

ABSTRACT

Neurocysticercosis (NC), i.e., the presence of the larval form of
Taenia solium
in tissues, is the most frequent and severe infection involving the central nervous system. Paired serum and cerebrospinal fluid (CSF) samples from patients with NC, CSF and serum samples from a control group, and serum samples from patients with other parasitoses were studied by enzyme-linked immunosorbent assay (ELISA) and by immunoblotting with
Taenia crassiceps
vesicular fluid antigen (Tcra) and
Taenia solium
total saline antigen (Tso) for the detection of immunoglobulin G antibodies. ELISAs carried out with the Tso and Tcra antigens showed 94.1 and 95.6% sensitivities, respectively, for the detection of antibodies in CSF and 70.6% and 91.2% sensitivities, respectively, for the detection of antibodies in serum, with 100% specificity for the detection of antibodies in CSF and 80% specificity for the detection of antibodies in serum for both antigens. On the basis of the reactivities of the peptides in the samples analyzed, the peptides of ≤23, 39, 85 to 77, and 97 kDa were found to be Tso specific by immunoblotting and the peptides of ≤62, 74, 109, 121, and 131 kDa were found to be Tcra specific. Tests with Tcra extract had higher sensitivities and more homogeneous results and permitted us to obtain the parasites easily. We suggest the use of Tcra ELISA for the study of serum and confirmation of the results for sera positive by an immunoblotting analysis in which specific peptides (e.g., peptides of 19 to 13 kDa) are detected.
Comparison of PCR-Restriction Fragment Length Polymorphism Analysis and PCR-Direct Sequencing Methods for Differentiating
Helicobacter pylori ureB
Gene Variants
Authors: Tanahashi Toshihito , Kita Masakazu , Kodama Tadashi , Sawai Naoki , Yamaoka Yoshio , Mitsufuji Shoji , Katoh Fumitaka , Imanishi Jiro 
Published: May 2021

ABSTRACT

A method utilizing PCR-restriction fragment length polymorphism (RFLP) in the
Helicobacter pylori
genes is widely used to differentiate strains. However, with this typing method only a single base change at a specific restriction site can be detected. In addition, it is unclear whether the nucleotide base change recognized by RFLP is related to a substitution of encoded amino acid. To examine the validity of the PCR-RFLP method, 933-bp PCR products were obtained from 41 different clinical
H. pylori
isolates and were digested with
Sau
3A restriction endonuclease. Furthermore, the nucleotides of the same region in the
ureB
gene were directly sequenced and compared. PCR-RFLP confirmed that there was genetic diversity within the
ureB
gene with three distinct types, one being well conserved and the other two being variations. However, the direct sequencing method revealed that there was no difference at the nucleotide level among these RFLP types. Base substitutions recognized by
Sau
3A occurred in the third-base position and did not change the encoded amino acid. In addition, many nucleotide mutations, which could not be recognized by
Sau
3A, were frequently found. These results suggest that the PCR-RFLP method provides for an easy typing scheme of isolates, but does not reveal the true extent of genetic diversity. It is proposed that careful observation is required for the interpretation of results when clinical isolates are differentiated.
Serodiagnosis of Recently Acquired
Toxoplasma gondii
Infection with a Recombinant Antigen
Authors: Li Shuli , Maine Greg , Suzuki Yasuhiro , Araujo Fausto G. , Galvan Gina , Remington Jack S. , Parmley Stephen 
Published: May 2021

ABSTRACT

A portion of a cDNA encoding a 35-kDa antigen from
Toxoplasma gondii
was cloned into the CKS expression vector and expressed in
Escherichia coli
. By using the enzyme-linked immunosorbent assay (ELISA), the recombinant protein (rP35 antigen) was examined for reactivity with immunoglobulin G (IgG) antibodies in the sera of pregnant women. Of these women, 41 had a toxoplasma serologic profile suggestive of recently acquired
T. gondii
infection (Sabin-Feldman dye test [DT] titers from 1:256 to 1:32,000, positive IgM ELISA titers from 2.3 to 9.7, positive IgA ELISA from 1 to >28, and acute patterns in the differential agglutination [AC/HS] test) (group I), and 50 women had a toxoplasma serologic profile suggestive of infection acquired in the distant past (low DT titers from 1:16 to 1:512, negative IgM ELISA titers from 0 to 0.8, and chronic patterns in the AC/HS test) (group II). The classification of acute or chronic profile was based on the individual's clinical history as well as the combination of the results of the toxoplasma serological profile. An additional group (group III) was composed of sera from 50 women who were seronegative for
T. gondii
antibodies in the DT. The results revealed that whereas 85.3% of women in group I had IgG antibodies that reacted with the rP35 antigen, only 8% of women in group II had IgG antibodies that reacted with the same antigen. In immunoblots, the rP35 antigen was recognized by IgG antibodies in a pool of sera from individuals with a toxoplasma serologic profile compatible with acute infection but not in a pool of sera from individuals with a serologic profile characteristic of a chronic infection. These results reveal that IgG antibodies against the P35 antigen are produced during the acute stage of the infection but are uncommon in the latent or chronic phase of the infection. Thus, the rP35 antigen may be a useful serologic marker to differentiate between recently acquired infection and that acquired in the more distant past.
Automated Specific Capture of Hepatitis C Virus RNA with Probes and Paramagnetic Particle Separation
Authors: Miyachi Hayato , Masukawa Atsuko , Ohshima Toshio , Hirose Toru , Impraim Chaka , Ando Yasuhiko 
Published: May 2021

ABSTRACT

We developed and evaluated a prototype automated specimen preparation instrument for the specific capture of hepatitis C virus (HCV) RNA with probes and magnetic bead-fluid separation. HCV RNA was isolated from serum by lysis of virus particles with a chaotropic agent, followed by hybridization of the RNA with biotinylated probes and capture of the hybridized RNA with streptavidin-coated paramagnetic particles. After washing of the hybrid-particle complexes to remove nonspecifically bound materials, the particles were resuspended in a specimen diluent and were then ready for amplification and detection with a fully automated PCR system (COBAS AMPLICOR; Roche Diagnostic Systems). The analytical sensitivity in the dilution series was 33 copies per ml or greater. Comparison of the test results with those obtained by a manual method based on organic extraction and precipitation of RNA (SepaGene RV-R; Sanko Junyaku Co., Ltd.) showed 93% (49 of 53 samples) sensitivity and 100% (12 of 12 samples) specificity. There was 94% overall agreement between results. When RNA was extracted by the manual method from serum containing 10
3
or 10
5
copies of HCV per ml in the presence of heparin, there was an inhibitory effect on detection of both HCV RNA and the internal control. In contrast, when RNA was extracted from the serum by the automated method, there was no inhibitory effect. This inhibitory effect of heparin on the manual method was also observed for a series of serum specimens from a hemodialysis patient, but the inhibitory effect was eliminated by the automated specimen preparation method. In summary, a fully automated RNA extraction system for PCR detection of HCV RNA by use of specific capture with probes and magnetic bead-fluid separation was shown to have performance similar to that of the conventional manual method. In addition, it successfully eliminated the inhibitory effect of the heparin in the serum and permitted the detection of HCV RNA in serum samples from a hemodialysis patient. The prototype automated RNA extraction system is suitable as a totally automated system, starting with RNA extraction to detection of HCV, if it was combined with the fully automated COBAS AMPLICOR PCR system.
Phenotypic and Molecular Typing of Nosocomial Methicillin-Resistant
Staphylococcus aureus
Strains Susceptible to Gentamicin Isolated in France from 1995 to 1997
Authors: Galdbart Jacques-Olivier , Morvan Anne , El Solh Nevine 
Published: May 2021

ABSTRACT

Methicillin-resistant strains susceptible to gentamicin (Gm
s
MRSA) have emerged since 1993 in several French hospitals. To study whether particular clones have spread in various French cities and whether some clones are related to gentamicin-resistant (Gm
r
) MRSA strains, various methods (antibiotyping, phage typing, determination of
Sma
I macrorestriction patterns before and after hybridization with IS
256
transposase and
aacA-aphD
probes) were used to compare 62 Gm
s
MRSA strains isolated from 1995 to 1997 in nine cities and 15 Gm
r
MRSA strains. Eighteen major
Sma
I genotypes were identified, of which 11 included only Gm
s
MRSA strains and 5 included only Gm
r
MRSA strains. Each of the Gm
r
MRSA strains contained 6 to 13
Sma
I fragments hybridizing with the insertion sequence IS
256
, of which a single band also hybridized with the
aacA-aphD
gene. No such hybridizing sequences were detected in 60 of the 62 Gm
s
MRSA strains. Thus, the divergence between Gm
r
and Gm
s
MRSA strains is revealed, not only by their distributions in distinct
Sma
I genotypes but also by the differences in hybridization patterns. Two of the 62 Gm
s
MRSA strains had the uncommon feature of carrying several
Sma
I bands hybridizing with IS
256
, suggesting that they are possibly related to the Gm
r
MRSA strains grouped in the same
Sma
I genotype. Five of the 11
Sma
I genotypes including only Gm
s
MRSA strains contained strains from diverse cities, isolated during different years and with different antibiograms, suggesting that some clones have spread beyond their cities of origin and persisted.
Restriction Fragment Length Polymorphism Analysis of
Mycobacterium tuberculosis
Isolated from Countries in the Western Pacific Region
Authors: Park Young-Kil , Bai Gill-Han , Kim Sang-Jae 
Published: May 2021

ABSTRACT

A total of 422
Mycobacterium tuberculosis
isolates from eight countries were subjected to IS
6110
and IS
1081
DNA fingerprinting by means of restriction fragment analysis to characterize
M. tuberculosis
strains from each country. Chinese, Mongolian, Hong Kong, Filipino, and Korean isolates had comparatively more copies of IS
6110
(proportion with eight or more copies; 95% ± 5%), while Thai, Malaysian, and Vietnamese isolates had fewer copies (proportion with eight or more copies, 60% ± 4%). We found a number of novel IS
1081
types in this study. One IS
1081
type was present in 56% of Filipino isolates, had a specific 6.6-kb
Pvu
II fragment in its IS
6110
DNA fingerprint, and was termed the “Filipino family.” The IS
1081
types of Thai isolates had interposing characteristics between the characteristics of northeastern Asian and southeastern Asian IS
1081
types. A 1.3-kb single-copy IS
6110
fragment was found only in Vietnamese
M. tuberculosis
isolates. Although
M. tuberculosis
isolates from each country had comparatively similar characteristics depending on the classification factor, each country's isolates showed characteristic DNA fingerprints and differed slightly from the isolates from the other countries in either the mode number of IS
6110
copies or the distribution of IS
1081
types.
The 1998 Senegal Epidemic of Meningitis Was Due to the Clonal Expansion of A:4:P1.9, Clone III-1, Sequence Type 5
Neisseria meningitidis
Strains
Authors: Nicolas Pierre , Raphenon Georges , Guibourdenche Martine , Decousset Laurent , Stor Richard , Gaye Abou Beckr 
Published: May 2021

ABSTRACT

Between January and April 1998, a meningitis outbreak due to serogroup A meningococcus took place in Senegal. The outbreak began in Gandiaye, 165 km to the east of Dakar, and progressed towards the towns of Gossas, Niakkhar, Guinguineo, Fatik, Foundiougne, Dioffior, Sokone, Kaolack, and Nioro. At the same time, the outbreak reached regions of Kaffrine, Koungheul, and Tambacounda in the east of Senegal. A total of 1,350 cases and 200 deaths were reported. The WHO Collaborating Center in Marseilles received 24 strains for analysis. All were serogroup A
Neisseria meningitidis
, type 4 and subtype P1.9. Multilocus enzyme electrophoresis, performed by Institut Pasteur Paris, showed that the strains belonged to clone III-1. DNA restriction fragments generated by endonuclease
Bgl
II and analyzed by pulsed-field gel electrophoresis showed 24 indistinguishable fingerprint patterns similar to those of meningococcus strains isolated from African outbreaks since 1988. Three strains were studied by multilocus sequence typing (MLST) with seven loci. The comparison between sequences and existing alleles on the MLST website (
http://mlst.zoo.ox.ac.uk
) allowed us to assign these strains to sequence type 5 (ST5), as their sequences were identical to the consensus at seven loci. All 24 strains were susceptible to penicillin, amoxicillin, chloramphenicol, and rifampin. Subgroup III is finishing its spread towards west of the meningitis belt of Africa. To our knowledge, this is the first time subgroup III, and more precisely ST5, strains are reported as being responsible for a meningitis outbreak in Senegal.
Detection of Clarithromycin-Resistant
Helicobacter pylori
Strains by a Preferential Homoduplex Formation Assay
Authors: Maeda Shin , Yoshida Haruhiko , Matsunaga Hironari , Ogura Keiji , Kawamata Osamu , Shiratori Yasushi , Omata Masao 
Published: May 2021

ABSTRACT

It has been shown that resistance to clarithromycin, a major cause of failure in
Helicobacter pylori
eradication therapy, is associated with point mutations in the 23S rRNA gene. We sought to apply the preferential homoduplex formation assay (PHFA), a novel technique for the efficient detection of point mutations, to detection of the mutations. PHFA was performed on streptavidin-coated microtiter plates with biotin- and dinitrophenyl-labeled amplicons to detect the wild-type gene or each mutant gene. DNA samples were extracted from gastric juice specimens of 412 patients with
H. pylori
infection and were applied to the assay. The detection threshold of PHFA was as few as 10 gene copies. The sensitivity of PHFA for the detection of
H. pylori
infection was higher than those of culture and the rapid urease test. A total of 337 (81.8%) samples had the wild-type gene, 38 (9.2%) had the A2144G mutation, and 37 (9.0%) contained both the wild type and a mutation (A2144G in 30 samples, A2143G in 5 samples, and A2143G plus A2144G in 2 samples). About half the strains isolated from patients with mixed infection were susceptible by the agar dilution method (MIC, <0.1 mg/liter). Therefore, PHFA can detect clarithromycin-resistant
H. pylori
strains, even in patients with mixed infections with the wild type, that are not detectable by the agar dilution method.
Comparative Analysis of PCR versus Culture for Diagnosis of Ulceroglandular Tularemia
Authors: Johansson Anders , Berglund Lennart , Eriksson Ulla , Göransson Ingela , Wollin Ralfh , Forsman Mats , Tärnvik Arne , Sjöstedt Anders 
Published: May 2021

ABSTRACT

PCR and culture were comparatively evaluated for their abilities to demonstrate
Francisella tularensis
in wound specimens from tularemia patients during an outbreak in Sweden in 1998. For transport of the specimens used for PCR, a buffer solution containing a nuclease inhibitor was used, and for transport of the specimens used for culture, a commercial transport system was selected after experimental comparison of various systems. Of 40 patients with culture- and/or serology-verified ulceroglandular tularemia, PCR detected
F. tularensis
DNA in 30 (75%) patients, whereas culture detected bacterial growth in 25 (62%) patients. Compared to data from a previous study, the present inclusion of a nuclease inhibitor in the transport medium did not improve the sensitivity of the PCR, whereas the sensitivity of the culture procedure was significantly increased by selection of the system used for transport. Among eight patients with clinically suspected tularemia but with negative serology and culture, specimens from four patients showed detectable DNA. In three of these patients the diagnosis was verified by the demonstration of an
F. tularensis
-specific T-cell response in vitro. In conclusion, PCR was more sensitive than culture for demonstration of
F. tularensis
in wound specimens. Besides, we showed that tularemia may proceed without development of serum antibodies, and in these patients, PCR may be of special importance for verification of the diagnosis.
Distribution of the Intermedilysin Gene among the Anginosus Group Streptococci and Correlation between Intermedilysin Production and Deep-Seated Infection with
Streptococcus intermedius
Authors: Nagamune Hideaki , Whiley Robert A. , Goto Takatsugu , Inai Yasuko , Maeda Takuya , Hardie Jeremy M. , Kourai Hiroki 
Published: May 2021

ABSTRACT

The distribution of intermedilysin, a human-specific cytolysin, among the anginosus group streptococci and the correlation of toxin production and infection by
Streptococcus intermedius
were investigated. PCR and Southern hybridization specific for the intermedilysin gene revealed that the toxin gene exists only in
S. intermedius
and no homologue to the toxin gene is distributed in
S. anginosus
and
S. constellatus
. Thus, the intermedilysin gene is useful as a marker gene of
S. intermedius
. Moreover, a human-specific hemolysis assay and Western blotting with intermedilysin-specific antibodies clearly demonstrated that the intermedilysin production level in isolates from deep-seated infections, such as brain and liver abscesses, is higher (6.2- to 10.2-fold, respectively) than in strains from normal habitats, such as dental plaque, or from peripheral infection sites. However, other candidate virulence factors of
S. intermedius
, such as chondroitin sulfate depolymerase, hyaluronidase, and sialidase activities, did not show such a clear correlation between enzymatic activity and isolation sites or disease severity. From these results, intermedilysin is likely to be the pathogenic or triggering factor of significance in inducing deep-seated infections with
S. intermedius
.
Typing of
Candida glabrata
in Clinical Isolates by Comparative Sequence Analysis of the Cytochrome
c
Oxidase Subunit 2 Gene Distinguishes Two Clusters of Strains Associated with Geographical Sequence Polymorphisms
Authors: Sanson Gerdine F. O. , Briones Marcelo R. S. 
Published: May 2021

ABSTRACT

We tested whether comparative sequence analysis of the mitochondrion-encoded cytochrome
c
oxidase subunit 2 gene (
COX2
) could be used to distinguish intraspecific variants of
Candida glabrata
. Mitochondrial genes are suitable for investigation of close phylogenetic relationships because they evolve much faster than nuclear genes, which in general exhibit very limited intraspecific variation. For this survey we used 11 clinical isolates of
C. glabrata
from three different geographical locations in Brazil, 10 isolates from one location in the United States, 1 American Type Culture Collection strain as an internal control, and the published sequence of strain CBS 138. The complete coding region of
COX2
was amplified from total cellular DNA, and both strands were sequenced twice for each strain. These sequences were aligned with published sequences from other fungi, and the numbers of substitutions and phylogenetic relationships were determined. Typing of these strains was done by using 17 substitutions, with 8 being nonsynonymous and 9 being synonymous. Also, cDNAs made from purified mitochondrial polyadenylated RNA were sequenced to confirm that our sequences correspond to the expressed copies and not nuclear pseudogenes and that a frameshift mutation exists in the 3′ end of the coding region (position 673) relative to the
Saccharomyces cerevisiae
sequence and the previously published
C. glabrata
sequence. We estimated the average evolutionary rate of
COX2
to be 11.4% sequence divergence/10
8
years and that phylogenetic relationships of yeasts based on these sequences are consistent with rRNA sequence data. Our analysis of
COX2
sequences enables typing of
C. glabrata
strains based on 13 haplotypes and suggests that positions 51 and 519 indicate a geographical polymorphism that discriminates strains isolated in the United States and strains isolated in Brazil. This provides for the first time a means of typing of
Candida
strains that cause infections by use of direct sequence comparisons and the associated divergence estimates.
Typing of
Histoplasma capsulatum
Isolates Based on Nucleotide Sequence Variation in the Internal Transcribed Spacer Regions of rRNA Genes
Authors: Jiang Bingdong , Bartlett Marilyn S. , Allen Stephen D. , Smith James W. , Wheat L. Joseph , Connolly Patricia A. , Lee Chao-Hung 
Published: May 2021

ABSTRACT

The nucleotide sequences of internal transcribed spacer (ITS) regions of rRNA genes of 24 isolates of
Histoplasma capsulatum
were examined. The results indicate that the sequences of ITS regions in different isolates are not identical. Sequence variations were found at 20 positions in the 496 bp that were sequenced. Ten different sequence patterns, designated types A through H, were observed when the sequences from the 24 isolates were aligned. Twelve isolates from Indianapolis were classified into four different types. Two isolates from New York belonged to type G. Three isolates from different cities were type F. The remaining six isolates were of different types.