bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evaluation of Protocol Using Gene Capture and PCR for Detection of
Helicobacter pylori
DNA in Feces
Authors: MacKay W. G. , Williams C. L. , McMillan M. , Ndip R. N. , Shepherd A. J. , Weaver L. T. 
Published: October 2003

ABSTRACT

The route of transmission of
Helicobacter pylori
, which is usually acquired in childhood and is one of the most common bacterial infections in humans, remains undetermined. Mapping the distribution of
H
.
pylori
genotypes within families could help to determine the routes of transmission and risk factors. Here we describe a noninvasive method for obtaining
H
.
pylori
DNA isolates from the feces of children. Children presenting with gastrointestinal symptoms at the Royal Hospital for Sick Children were tested for gastric
H
.
pylori
colonization by using the
13
C-urea breath test (UBT) and were asked to provide fecal samples, which were tested for
H
.
pylori
by using the HpSA fecal antigen test. DNA was purified from fecal samples by using a novel method of gene capture with subsequent
H
.
pylori
PCR analysis. Fifteen UBT-positive and 15 UBT-negative children participated in the study. The positive and negative predictive values for the assay were 80 and 100%, respectively. Fecal DNA purification followed by
H
.
pylori
PCR analysis is an effective tool for harvesting
H
.
pylori
DNA isolates from the feces of children. This technique may be developed to allow the diagnosis and noninvasive genotyping of
H
.
pylori
in children and their families.
Genome Type Analysis of Adenovirus Types 3 and 7 Isolated during Successive Outbreaks of Lower Respiratory Tract Infections in Children
Authors: Kim Yae-Jean , Hong Jung-Yun , Lee Hoan-Jong , Shin Seon-Hee , Kim Yun-Kyung , Inada Toshiki , Hashido Madoka , Piedra Pedro A. 
Published: October 2003

ABSTRACT

Adenovirus is an important cause of respiratory infections in infants and children. Fifty-one serotypes have been identified, and adenovirus type 3 (Ad3) and Ad7 have often been associated with outbreaks of severe respiratory tract infections. Each serotype can be further divided into genome types based on the patterns of digestion of their DNAs with restriction enzymes. DNA restriction analysis was performed with 56 strains of Ad3 and 98 strains of Ad7 by using 12 restriction enzymes recognizing 6 bp (
Bam
HI,
Bcl
I,
Bgl
I,
Bgl
II,
Bst
EII,
Eco
RI,
Hin
dIII,
Hpa
I,
Sal
I,
Sma
I,
Xba
I, and
Xho
I). The virus strains were isolated during outbreaks of lower respiratory tract infections in children during an 11-year period from 1990 to 2000 in Seoul, Korea. Among the Ad3 strains, seven genome types were identified; Ad3a and six novel types (Ad3a13, Ad3a14, Ad3a15, Ad3a16, Ad3a17, and Ad3a18). Multiple genome types cocirculated during outbreaks, and some of these were isolated during the 11-year observation period, while others were restricted to particular outbreaks. For Ad7, two genome types, Ad7d and Ad7l, the latter of which is a novel genome type, were identified. A shift in genome types occurred from Ad7d to Ad7l during successive outbreaks. Mortality was 3.6% among children with Ad3 infections and 18% among children infected with either of the Ad7 genome types. In conclusion, the data confirm that Ad3 genome types are more diverse than those of Ad7 and suggest that shifts of genome types may occur during successive outbreaks of Ad3 and Ad7.
Segregation of Human Immunodeficiency Virus Type 1 Subtypes by Risk Factor in Australia
Authors: Herring Belinda L. , Ge Ying C. , Wang Bin , Ratnamohan Mala , Zheng Frank , Cunningham Anthony L. , Saksena Nitin K. , Dwyer Dominic E. 
Published: October 2003

ABSTRACT

The aim of this study was to determine which human immunodeficiency virus type 1 (HIV-1) subtypes were circulating in Australia and to correlate the subtypes with risk factors associated with the acquisition of HIV-1 infection. DNA was extracted from peripheral blood mononuclear cells, and HIV-1
env
genes were amplified and subtyped using heteroduplex mobility analysis, with selected samples sequenced and phylogenetic analysis performed. The HIV-1
env
subtypes were determined for 141 samples, of which 40 were from female patients and 101 were from male patients; 13 samples were from children. Forty-seven patients were infected by homosexual or bisexual contact, 46 were infected through heterosexual contact, 21 were infected from injecting drug use (IDU), 13 were infected by vertical transmission, 8 were infected from nosocomial exposure, and 6 were infected by other modes of transmission, including exposure to blood products, ritualistic practices, and two cases of intrafamilial transmission. Five subtypes were detected; B (
n
= 104), A (
n
= 5), C (
n
= 17), E (CRF01_AE;
n
= 13), and G (
n
= 2). Subtype B predominated in HIV-1 acquired homosexually (94% of cases) and by IDU (100%), whereas non-subtype B infections were mostly seen in heterosexually (57%) or vertically (22%) acquired HIV-1 infections and were usually imported from Africa and Asia. Subtype B strains of group M viruses predominate in Australia in HIV-1 transmitted by homosexual or bisexual contact and IDU. However, non-B subtypes have been introduced, mostly acquired via heterosexual contact.
Molecular Evidence of Persistent Echovirus 13 Meningoencephalitis in a Patient with Relapsed Lymphoma after an Outbreak of Meningitis in 2000
Authors: Archimbaud Christine , Bailly Jean-Luc , Chambon Martine , Tournilhac Olivier , Travade Philippe , Peigue-Lafeuille Hélène 
Published: October 2003

ABSTRACTEnteroviral meningoencephalitis was diagnosed in a patient with an immunodeficiency syndrome acquired after treatment with rituximab for a relapsed primary B-cell lymphoma. A second meningoencephalitic episode was diagnosed 6 months later and was successfully treated with a combination of immunoglobulins and pleconaril. The infection was persistent since the enterovirus genome was detected in five sequential specimens of cerebrospinal fluid collected over 9 months. An echovirus 13 isolate was isolated in the first three samples. The viral sequence encoding the VP1 capsid protein of the three isolates was determined and was compared with that of four control viruses. The virus isolates recovered from the patient shared >99% nucleotide sequence similarity with one another. In a phylogenetic tree, they were directly related to a control virus obtained from a patient hospitalized in 2000 during an outbreak of enterovirus meningitis. The epidemiological origin of a chronic echovirus infection in a patient with immune deficiency suggests that the echovirus had been continuously circulating in the general population after the outbreak that had revealed its emergence.
Predominance ofEhrlichia ewingiiin Missouri Dogs
Authors: Liddell Allison M. , Stockham Steven L. , Scott Michael A. , Sumner John W. , Paddock Christopher D. , Gaudreault-Keener Monique , Arens Max Q. , Storch Gregory A. 
Published: October 2003

ABSTRACTTo investigate the species distribution ofEhrlichiapresent in Missouri dogs, we tested 78 dogs suspected of having acute ehrlichiosis and 10 healthy dogs. Blood from each dog was screened with a broad-range 16S rRNA gene PCR assay that detects known pathogenic species ofEhrlichiaandAnaplasma. The species was determined by using species-specific PCR assays and nucleotide sequencing.Ehrlichiaantibody testing was performed by using an indirect immunofluorescence assay withEhrlichia chaffeensisas the antigenic substrate. The broad-range assay detectedEhrlichiaorAnaplasmaDNA in 20 (26%) of the symptomatic dogs and 2 (20%) of the asymptomatic dogs.E. ewingiiaccounted for 20 (91%), andE. chaffeensisaccounted for 1 (5%) of the positives.Anaplasma phagocytophilumDNA was detected in one dog, and the sequences of regions of the 16S rRNA gene and thegroESLoperon amplified from the blood of this dog matched the published sequences of this organism. Antibodies reactive withE. chaffeensiswere detected in 14 (67%) of the 21 PCR-positive dogs and in 12 (19%) of the 64 PCR-negative dogs. Combining the results of PCR and serology indicated that 33 (39%) of 85 evaluable dogs had evidence of past or currentEhrlichiainfection. We conclude thatE. ewingiiis the predominant etiologic agent of canine ehrlichiosis in the areas of Missouri included in this survey.E. canis, a widely recognized agent of canine ehrlichiosis, was not detected in any animal. The finding ofE. ewingiiin asymptomatic dogs suggests that dogs could be a reservoir for thisEhrlichiaspecies.
Evaluation of the Hodge Test and the Imipenem-EDTA Double-Disk Synergy Test for Differentiating Metallo-β-Lactamase-Producing Isolates of
Pseudomonas
spp. and
Acinetobacter
spp
Authors: Lee K. , Lim Y. S. , Yong D. , Yum J. H. , Chong Y. 
Published: October 2003

ABSTRACT
Gram-negative bacilli with acquired metallo-β-lactamase (MBL) production have been increasingly reported in some countries, necessitating their detection. The aim of this study was to evaluate the performance of the Hodge test and those of the imipenem (IPM)-EDTA, ceftazidime (CAZ)-mercaptopropionic acid (MPA), and CAZ-sodium mercaptoacetic acid (SMA) double-disk synergy tests (DDSTs). The efficiencies of testing CAZ-resistant and IPM-nonsusceptible isolates were also compared. Strains used for the evaluation were known IMP-1 and VIM-2 MBL-producing isolates and consecutive and CAZ-nonsusceptible isolates of pseudomonads and acinetobacters. The performance of the Hodge test was improved by addition of zinc sulfate (140 μg/disk) to an IPM disk. In DDSTs, EDTA (ca. 1,900 μg) disks were better at detecting MBL-producing strains among pseudomonads, while MPA (3 μl) and SMA (3 mg) disks performed better for acinetobacters. EDTA (ca. 750 μg)-plus-SMA (ca. 2 mg) disks performed better than EDTA, MPA, or SMA disks with both organisms. CAZ-SMA DDSTs failed to detect 22 of 80 (28%) MBL-producing acinetobacters. In conclusion, use of an IPM disk and an EDTA (750 μg)-plus-SMA (2 mg) disk improves performance, and testing IPM-nonsusceptible isolates rather than CAZ-resistant isolates could reduce screening work. Further evaluation of the test is required for the detection of other types of MBL-producing gram-negative bacilli.
Detection of a Point Mutation Associated with High-Level Isoniazid Resistance in
Mycobacterium tuberculosis
by Using Real-Time PCR Technology with 3′-Minor Groove Binder-DNA Probes
Authors: van Doorn H. Rogier , Claas Eric C. J. , Templeton Kate E. , van der Zanden Adri G. M. , te Koppele Vije Arianne , de Jong Menno D. , Dankert Jacob , Kuijper Ed J. 
Published: October 2003

ABSTRACT

Tuberculosis remains one of the leading infectious causes of death worldwide. The emergence of drug-resistant strains of
Mycobacterium tuberculosis
is a serious public health threat. Resistance to isoniazid (INH) is the most prevalent form of resistance in
M. tuberculosis
and is mainly caused by mutations in the catalase peroxidase gene (
katG
). Among high-level INH-resistant isolates (MIC ≥ 2), 89% are associated with a mutation at codon 315 of
katG
. There is a need to develop rapid diagnostic tests to permit appropriate antibiotic treatment and to improve clinical management. Therefore, a single-tube real-time PCR, using a novel kind of probe (3′-minor groove binder-DNA probe), was developed to detect either the wild-type or the mutant codon directly in Ziehl-Neelsen-positive sputum samples. The detection limit of the assay for purified DNA was 5 fg per well (one mycobacterial genome), and with spiked sputum samples, it was 20 copies per well, corresponding to 10
3
mycobacteria per ml of sputum. Sputum samples from 20 patients living in Kazakhstan or Moldova and infected with monodrug- or multidrug-resistant
M. tuberculosis
and 20 sputum samples from patients infected with INH-susceptible
M. tuberculosis
were tested. The sensitivities and specificities of the probes were 70 and 94% for the wild-type probe and 82 and 100% for the mutant probe. Binding to either probe was nonambiguous. This real-time PCR allows the rapid identification of a mutant
katG
allele and can easily be implemented in a clinical microbiology laboratory.
Multiplexed, Real-Time PCR for Quantitative Detection of Human Adenovirus
Authors: Gu Z. , Belzer S. W. , Gibson C. S. , Bankowski M. J. , Hayden R. T. 
Published: October 2003

ABSTRACT

Adenovirus infection is becoming increasingly recognized as a cause of morbidity and mortality in the immunosuppressed patient population. While early detection and quantitation of adenovirus in peripheral blood has been suggested as a means of directing and monitoring antiviral therapy in these patients, few methods have been published, particularly with respect to viral quantitation. A multiplexed real-time PCR assay was developed that can quantitatively detect a wide range of known serotypes of human adenovirus, including all of subgroups A to C. This assay was compared to a qualitative, Southern blot-based PCR assay by using 45 peripheral blood specimens from 16 patients. There was 100% concordance between the two tests in terms of qualitative results. The real-time assay detected adenovirus in patient samples at levels from <200 to 266,681 copies/ml of blood. By using control viral samples, sensitivity was demonstrated to less than 10 copies of viral genome per reaction and quantitative linearity was demonstrated from 10 to 10
6
copies of input viral DNA. Equivalent sensitivity and linearity were demonstrated for 15 different reference serotypes of adenovirus. Eleven other viral serotypes have complete target region sequence homology to one or more of the strains tested. No cross-reactivity was noted with other commonly isolated viral species. Sequence analysis showed no significant homology with any other human pathogens (bacterial or viral). This assay allows rapid, sensitive, and specific quantitation of adenovirus and may have a significant impact on the care of immunocompromised patients at risk for disseminated viral infection.
Human Metapneumovirus Infection in the Canadian Population
Authors: Bastien Nathalie , Ward Diane , Van Caeseele Paul , Brandt Ken , Lee Spencer H. S. , McNabb Gail , Klisko Brian , Chan Edward , Li Yan 
Published: October 2003

ABSTRACT

Human metapneumovirus (hMPV), a newly discovered paramyxovirus, has been associated with acute respiratory tract infections (ARIs) ranging from upper ARIs to severe bronchiolitis and pneumonia. Important questions remain on the contribution of hMPV to ARIs and its impact on public health. During the 2001-2002 season, we conducted a collaborative study with four provincial public health laboratories to study the prevalence of this new virus in the Canadian population. A total of 445 specimens were collected from patients of all age groups with ARIs and were tested for the presence of hMPV by reverse transcription-PCR. Of these, 66 (14.8%) tested positive for hMPV. Positive specimens were found in all age groups and in all four provinces studied. Virus activity peaked in February and March. The age range of the patients with hMPV infection was 2 months to 93 years (median age, 25 years), with similar numbers of females (35%) and males (41%). Thirty-three percent (
n
= 22) of hMPV-infected patients were hospitalized; of these, 27% (
n
= 6) had rhinitis and pneumonia, 23% (
n
= 5) had bronchiolitis, and 9% (
n
= 2) had bronchitis. The hospitalization rates were significantly higher among patients <5 years of age (
P
= 0.0005) and those >50 years of age (
P
= 0.0044) than among those 6 to 50 years of age. Phylogenetic analysis of the F gene showed that two hMPV genetic clusters were cocirculating in the 2001-2002 season, and comparison with earlier studies suggests a temporal evolutionary pattern of hMPV isolates. These results provide further evidence of the importance of hMPV in ARIs, particularly in young children and elderly individuals.
Use of 16S rRNA Gene Sequencing for Rapid Identification and Differentiation ofBurkholderia pseudomalleiandB. mallei
Authors: Gee Jay E. , Sacchi Claudio T. , Glass Mindy B. , De Barun K. , Weyant Robbin S. , Levett Paul N. , Whitney Anne M. , Hoffmaster Alex R. , Popovic Tanja 
Published: October 2003

ABSTRACTBurkholderia pseudomalleiandB. mallei, the causative agents of melioidosis and glanders, respectively, are designated category B biothreat agents. Current methods for identifying these organisms rely on their phenotypic characteristics and an extensive set of biochemical reactions. We evaluated the use of 16S rRNA gene sequencing to rapidly identify these two species and differentiate them from each other as well as from closely related species and genera such asPandoraeaspp.,Ralstoniaspp.,Burkholderia gladioli,Burkholderia cepacia,Burkholderia thailandensis, andPseudomonas aeruginosa. We sequenced the 1.5-kb 16S rRNA gene of 56B. pseudomalleiand 23B. malleiisolates selected to represent a wide range of temporal, geographic, and origin diversity. Among all 79 isolates, a total of 11 16S types were found based on eight positions of difference. Nine 16S types were identified inB. pseudomalleiisolates based on six positions of difference, with differences ranging from 0.5 to 1.5 bp. Twenty-two of 23B. malleiisolates showed 16S rRNA gene sequence identity and were designated 16S type 10, whereas the remaining isolate was designated type 11. This report provides a basis for rapidly identifying and differentiatingB. pseudomalleiandB. malleiby molecular methods.
Detection of
Staphylococcus aureus
Enterotoxins A to D by Real-Time Fluorescence PCR Assay
Authors: Klotz M. , Opper S. , Heeg K. , Zimmermann S. 
Published: October 2003

ABSTRACT

Staphylococcus aureus
is one of the most significant pathogens causing nosocomial and community-acquired infections. Among the secreted staphylococcal virulence factors, there is a growing list of enterotoxins which can induce gastroenteric syndrome and toxic shock syndrome. Here, we developed a real-time fluorescence PCR assay (TaqMan PCR) for the detection of genes encoding staphylococcal enterotoxins A, B, C1, and D (SEA, SEB, SEC1, and SED) of
S
.
aureus
as well as the
mecA
gene encoding methicillin resistance and the
femB
gene as a specific genomic marker for
S
.
aureus
. SEA to SED were selected because they are the four classically described enterotoxins of
S
.
aureus
and because they were detected by latex agglutination. In order to evaluate the reliability of TaqMan PCR, we investigated 93 isolates of
S
.
aureus
derived from patients at our hospital over 5 months and compared the results with data obtained by a commerciall available reversed passive latex agglutination assay (SET-RPLA) for these isolates. Thirteen enterotoxin genes were detected by TaqMan PCR; however, no proteins expressed by these genes were detected by SET-RPLA. As a result, more isolates of
S
.
aureus
(
n
= 44) were found positive by TaqMan PCR for one or more enterotoxin genes than by SET-RPLA for the respective proteins expressed by these genes (
n
= 40). We conclude that TaqMan PCR is more sensitive because it offers the possibility for determining enterotoxins on a genotypic basis. Additionally, the assay allows the parallel detection of genes for SEA to SED and methicillin resistance in
S
.
aureus
. Furthermore, real-time PCR is well suited for screening large numbers of samples at the same time, allowing rapid, reliable, efficient, and cost-saving routine laboratory diagnosis.
Six-Month Multicenter Study on Invasive Infections Due to Group B Streptococci in Argentina
Authors: Lopardo Horacio A. , Vidal Patricia , Jeric Paola , Centron Daniela , Paganini Hugo , Facklam Richard R. , Elliott John 
Published: October 2003

ABSTRACT

There is little information about invasive infections by group B streptococci (GBS) and their antimicrobial susceptibilities in Latin America. We performed a prospective multicenter study to determine the serotype distribution and the antimicrobial susceptibility of GBS in Argentina. We identified 58 cases, but only 44 had sufficient data to be evaluated. Eight early-, four late-, and one fatal late, late-onset neonatal infections due to GBS were found. A total of 31 patients were adults with bacteremia, skin and soft tissue infections, osteomyelitis, arthritis, meningitis, abdominal infections, and renal abscess. Serotype III was prevalent in late-onset neonatal disease, and several serotypes (Ia/c, III, Ia, and II) were involved in early-onset neonatal infections. Serotypes II, Ia/c, III, and IV were commonly found in adults, with serotype II prevalent in younger adults (18 to 69 years old) and serotype Ia/c prevalent in elderly adults (>70 years old). The mortality rate attributable to GBS infections was 10.8%. All GBS were susceptible to penicillin and ceftriaxone. Resistance to clindamycin (1.7%), erythromycin (5.2%), azithromycin (5.2%), minocycline (69%), and tetracycline (72.4%), to high levels of kanamycin and amikacin (1.7%), and to intermediately high levels of gentamicin (1.7%) was observed. The bifunctional enzyme AAC6′-APH2" was detected in the isolate resistant to aminoglycosides, and other genetic determinants were identified in other resistant isolates:
tetM
and
tetO
in tetracycline-resistant streptococci and
mefA
and
ermTR
for efflux-mediated and inducible macrolide-lincosamide-streptogramin B-resistant streptococci, respectively. For clinical purposes and rapid and easy detection of high-level aminoglycoside-resistant GBS, a screening method that used 1,000-μg kanamycin disks is proposed.
Use of Turbidimetric Growth Curves for Early Determination of Antifungal Drug Resistance of Filamentous Fungi
Authors: Meletiadis Joseph , te Dorsthorst Debbie T. A. , Verweij Paul E. 
Published: October 2003

ABSTRACT

A previously described microbroth kinetic system (J. Meletiadis, J. F. Meis, J. W. Mouton, and P. E. Verweij, J. Clin. Microbiol.
39:
478-484, 2001) based on continuous monitoring of changes in the optical density of fungal growth was used to describe turbidimetric growth curves of different filamentous fungi in the presence of increasing concentrations of antifungal drugs. Therefore, 24 clinical mold isolates, including
Rhizopus oryzae
,
Aspergillus fumigatus
,
Aspergillus flavus
, and
Scedosporium prolificans
, were tested against itraconazole, terbinafine, and amphotericin B according to NCCLS guidelines. Among various parameters of the growth curves, the duration of the lag phase was strongly affected by the presence of antifungal drugs. Exposure to increasing drug concentrations resulted in prolonged lag phases of the turbidimetric growth curves. The lag phases of the growth curves at drug concentrations which resulted in more than 50% growth (for itraconazole and terbinafine) and more than 75% growth (for amphotericin B) after 24 h of incubation for
R. oryzae
, 48 h for
Aspergillus
spp., and 72 h for
S. prolificans
were 4 h longer than the lag phases of the growth curves at the corresponding drug-free growth controls which varied from 4.4 h for
R. oryzae
, 6.5 h for
A. flavus
, 7.9 h for
A. fumigatus
, and 11.6 h for
S. prolificans
. The duration of the lag phases showed small experimental and interstrain variability, with differences of less than 2 h in most of the cases. Using this system, itraconazole and terbinafine resistance (presence of >50% growth) as well as amphotericin B resistance (presence of >75% growth) was determined within incubation periods of 5.0 to 7.7 h for
R. oryzae
(for amphotericin B resistance incubation for up to 12 h was required), 8.8 to 11.4 h for
A. fumigatus
, 6.7 to 8.5 h for
A. flavus
, and 13 to 15.6 h for
S. prolificans
while awaiting formal MIC determination by the NCCLS reference method.