bluuebunny/crossref_metadata_2025_split huggingface dataset:
Discrimination within Phenotypically Closely Related Definitive Types of
Salmonella enterica
Serovar Typhimurium by the Multiple Amplification of Phage Locus Typing Technique
Authors: Ross Ian L. , Heuzenroeder Michael W. 
Published: April 2005

ABSTRACT

Multilocus sequence typing (MLST) is a relatively new high-resolution typing system employed for epidemiological studies of bacteria, including
Salmonella
. Discrimination based on MLST of housekeeping genes may be problematical, due to the high identity of gene sequences of closely related
Salmonella
species. The presence of genomic sequences derived from stable temperate phages in
Salmonella
offers an alternative for MLST of
Salmonella
. We have used MLST of prophage loci in
Salmonella enterica
serovar Typhimurium to discriminate closely related isolates of serovar Typhimurium. We have compared these results to MLST of five housekeeping genes, as well as pulsed-field gel electrophoresis (PFGE). The presence or absence of prophage loci in the 73 serovar Typhimurium isolates tested, as well as allelic variation as detected by sequencing, provided greater discrimination between isolates than either MLST of housekeeping genes or PFGE. Amplification of prophage loci alone separated serovar Typhimurium isolates into 27 groups comprising multiple isolates or individual strains. Sequencing of isolates found within the clusters separated isolates even further. By contrast, PFGE could only divide the 73 isolates into five distinct groups. MLST using housekeeping genes did not provide any significant separation of isolates in comparison to amplification or MLST of prophage loci. The results demonstrate that the amplification and sequencing of prophage loci provides a high-resolution, objective method for the discrimination of closely related isolates of serovar Typhimurium. It is proposed that multiple amplification of phage locus typing may provide sufficient discrimination for epidemiological purposes without recourse to MLST.
Rapid Diagnostic Method for Detection of Mumps Virus Genome by Loop-Mediated Isothermal Amplification
Authors: Okafuji Takao , Yoshida Naoko , Fujino Motoko , Motegi Yoshie , Ihara Toshiaki , Ota Yoshinori , Notomi Tsugunori , Nakayama Tetsuo 
Published: April 2005

ABSTRACT
Most mumps patients are clinically diagnosed without any virological examinations, but some diagnosed cases of mumps may be caused by other pathogens or secondary vaccine failure (SVF). To clarify these issues, a sensitive, specific, and rapid diagnostic method is required. We obtained 60 salivary swabs from 34 patients with natural infection during the course of the illness, 10 samples from patients with vaccine-associated parotitis, and 5 samples from patients with SVF. Total RNA was extracted and subjected to reverse transcription-PCR (RT-PCR) and loop-mediated isothermal amplification (LAMP) for genome amplification. We detected mumps virus RNA corresponding to 0.1 PFU by LAMP within 60 min after RNA extraction, with the same sensitivity as RT-nested PCR. Mumps virus was isolated in 30 of 33 samples within day 2, and mumps virus genome was amplified by LAMP in 32 of them. The quantity of virus titer was calculated by monitoring the time to reach the threshold of turbidity. The viral load decreased after day 3 and was lower in patients serologically diagnosed as having SVF with milder illness. Accuracy of LAMP for the detection of mumps virus genome was confirmed; furthermore, it is of benefit for calculating the viral load, which reflects disease pathogenesis.
Utility of Pooled Urine Specimens for Detection of
Chlamydia trachomatis
and
Neisseria gonorrhoeae
in Men Attending Public Sexually Transmitted Infection Clinics in Mumbai, India, by PCR
Authors: Lindan Christina , Mathur Meenakshi , Kumta Sameer , Jerajani Hermangi , Gogate Alka , Schachter Julius , Moncada Jeanne 
Published: April 2005

ABSTRACT

Pooling urogenital specimens for the detection of
Chlamydia trachomatis
and
Neisseria gonorrhoeae
by nucleic acid amplification tests is an attractive alternative to individual testing. As pooling can reduce the costs of testing as well as labor, it has been advocated for use in resource-poor settings. However, it has neither been widely adopted nor evaluated for use in developing countries. We evaluated the practical use of pooling first-catch urine (FCU) specimens for the detection of
C. trachomatis
and
N. gonorrhoeae
from 690 men in Mumbai, India, by PCR. FCU, urethral smears, and swabs were collected from men seen at two sexually transmitted infection (STI) clinics. All laboratory testing was done at the Lokmanya Tilak General Hospital. Gram stain smears and culture isolation for
N. gonorrhoeae
were performed. Each FCU was tested individually and in pools using the Roche Amplicor PCR for
C. trachomatis
and
N. gonorrhoeae
with an internal control for inhibition. Specimen pools consisted of aliquots from five consecutively processed FCUs combined into an amplification tube. An optical density reading of ≥0.20 indicated a pool for which subsequent testing of individual samples was required. Prevalence by PCR on single specimens was 2.2% (15/690) for
C. trachomatis
and 5.4% (37/690) for
N. gonorrhoeae
. Compared to individual FCU results, pooling for
C. trachomatis
and
N. gonorrhoeae
had an overall sensitivity of 96.1% (50/52). Specificity was 96.5% (83/86) in that three pools required single testing that failed to identify a positive specimen. Pooling missed two positive specimens, decreased the inhibition rate, and saved 50.3% of reagent costs. In this resource-limited setting, the use of pooling to detect
C. trachomatis
and
N. gonorrhoeae
by PCR proved to be a simple, accurate, and cost-effective procedure compared to individual testing.
Invasive Group A Streptococcal Disease in Alberta, Canada (2000 to 2002)
Authors: Tyrrell Gregory J. , Lovgren Marguerite , Kress Bertha , Grimsrud Karen 
Published: April 2005

ABSTRACT
Invasive group A streptococcal (iGAS) disease was placed under surveillance in Alberta in August 1999. The purpose of this study was to determine the incidence rates of iGAS infections throughout Alberta over a 3-year period (2000 to 2002) and to better understand the epidemiology of iGAS in this province. There were a total of 441 cases of invasive GAS disease over the 3 years examined (average population over 3 years, 3,055,765) and 47 deaths. The incidence in Alberta was 5.0 (2000), 5.7 (2001), and 3.8 (2002) per 100,000. The two main metropolitan regions (Edmonton and Calgary) had the majority of iGAS disease cases (305 cases), producing incidence rates of 4.8 (Edmonton) and 6.9 (Calgary) in 2000, 6.9 (Edmonton) and 6.6 (Calgary) in 2001, and 4.1 (Edmonton) and 3.9 (Calgary) in 2002, as well as deaths attributable to GAS (31 deaths). The three most prevalent M types were M1 (71 cases), M3 (52 cases), and MPT2967 (44 cases). With respect to age, the highest incidence rates occurred in those less than 1 year old (11.7 per 100,000) and those 65 years or older (11.5 per 100,000). Varicella virus infection preceded iGAS disease in 25% of children 8 years of age and under. A seasonal association was observed during the 3 years studied, with the highest number of cases occurring in the winter months and the lowest occurring during the summer months. The data for years 2000 and 2001 show that the metropolitan regions of Alberta experienced some of the highest incidence rates reported in North America in the past decade.
Use of Variable-Number Tandem Repeats To Examine Genetic Diversity of
Neisseria meningitidis
Authors: Yazdankhah Siamak P. , Lindstedt Bjørn-Arne , Caugant Dominique A. 
Published: April 2005

ABSTRACT

Repetitive DNA motifs with potential variable-number tandem repeats (VNTR) were identified in the genome of
Neisseria meningitidis
and used to develop a typing method. A total of 146 meningococcal isolates recovered from carriers and patients were studied. These included 82 of the 107
N. meningitidis
isolates previously used in the development of multilocus sequence typing (MLST), 45 isolates recovered from different counties in Norway in connection with local outbreaks, and 19 serogroup W135 isolates of sequence type 11 (ST-11), which were recovered in several parts of the world. The latter group comprised isolates related to the Hajj outbreak of 2000 and isolates recovered from outbreaks in Burkina Faso in 2001 and 2002. All isolates had been characterized previously by MLST or multilocus enzyme electrophoresis (MLEE). VNTR analysis showed that meningococcal isolates with similar MLST or MLEE types recovered from epidemiologically linked cases in a defined geographical area often presented similar VNTR patterns while isolates of the same MLST or MLEE types without an obvious epidemiological link showed variable VNTR patterns. Thus, VNTR analysis may be used for fine typing of meningococcal isolates after MLST or MLEE typing. The method might be especially valuable for differentiating among ST-11 strains, as shown by the VNTR analyses of serogroup W135 ST-11 meningococcal isolates recovered since the mid-1990s.
Characterization of the Genetic Lineages Responsible for Pneumococcal Invasive Disease in Portugal
Authors: Serrano I. , Melo-Cristino J. , Carriço J. A. , Ramirez M. 
Published: April 2005

ABSTRACT

The availability of a conjugate vaccine has the potential to reduce the disease burden of pneumococci and to alter the serotype frequency in the disease-causing population through immunoselection. These changes will probably be reflected in the distributions of individual genetic lineages within the population. We present a characterization of a collection of recent (1999 to 2002) invasive isolates from Portugal (
n
= 465) by macrorestriction profiling with pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. During this time, serotypes 14, 1, 3, 4, 8, 9V, 23F, 7F, 19A, and 12B were the 10 most prevalent overall by decreasing rank order. By combining the PFGE data with the sequence types (STs) of 104 isolates, we were able to identify the genetic lineages of the majority of the isolates. We found 66 STs, including 20 novel STs, corresponding to 47 different lineages by e-BURST analysis. We found in our collection a number of previously identified internationally disseminated lineages, especially among macrolide-resistant and penicillin-resistant isolates, and these accounted for most of the isolates. Most of the major lineages (17 of 25) were identified in all years of the study, suggesting that the pneumococcal population associated with invasive disease was stable. This study provides a characterization of the pneumococcal population associated with invasive disease that will be useful for detecting potential selective effects of the novel conjugate vaccine.
Testing for Induction of Clindamycin Resistance in Erythromycin-Resistant Isolates of
Staphylococcus aureus
Authors: Steward Christine D. , Raney Patti M. , Morrell Allison K. , Williams Portia P. , McDougal Linda K. , Jevitt Laura , McGowan John E. , Tenover Fred C. 
Published: April 2005

ABSTRACT

Disk diffusion and broth microdilution (BMD) were used to perform clindamycin (CLI) induction testing on 128 selected nonduplicate isolates of
Staphylococcus aureus
. Disk diffusion testing involved placing CLI and erythromycin (ERY) disks approximately 12 mm apart (measured edge to edge) on a Mueller-Hinton agar plate that had been inoculated with an
S. aureus
isolate; the plate was then incubated for 16 to 18 h. Two distinct induction phenotypes (labeled D and D
+
) and four noninduction phenotypes (designated as negative [Neg], hazy D zone [HD], resistant [R], and susceptible [S]) were observed in disk diffusion results. A clear, D-shaped zone of inhibition around the CLI disk was designated as the D phenotype and was observed for 21 isolates while a D-shaped zone containing inner colonies growing up to the CLI disk was designated as D
+
(17 isolates). In addition, 10 isolates were CLI susceptible and ERY resistant but were not inducible and showed no blunting of the CLI zone (Neg phenotype). Isolates that were CLI and ERY resistant (constitutive macrolide-lincosamide-streptogramin B resistance) demonstrated either a double zone of inhibition with an inner ring of reduced growth up to the edge of the disks (HD phenotype; 33 isolates) or solid growth around the CLI and ERY disks (R phenotype; 16 isolates). Finally, 31 isolates were susceptible by disk testing to both CLI and ERY (S phenotype). PCR results showed that isolates with a D phenotype harbored
ermA
, isolates with a D
+
phenotype contained either
ermC
(16 isolates) or
ermA
and
ermC
(one isolate), and all 10 isolates with a Neg phenotype contained
msrA
. All isolates with an HD or R phenotype harbored at least one
erm
gene. Isolates showing the D
+
phenotype by disk diffusion were also detected by BMD using a variety of CLI and ERY concentrations; however, isolates with the D phenotype were more difficult to detect by BMD and will likely require optimization of ERY and CLI concentrations in multilaboratory studies to ensure adequate sensitivity. Thus, at present, disk diffusion is the preferred method for testing
S. aureus
isolates for inducible CLI resistance.
CHROMagar Candida Medium for Direct Susceptibility Testing of Yeast from Blood Cultures
Authors: Tan Grace L. , Peterson Ellena M. 
Published: April 2005

ABSTRACT

An evaluation was performed on 95 blood cultures positive for
Candida
spp. to determine the correlation of direct susceptibility testing of fluconazole versus both standardized disk diffusion and MIC methods. For direct testing, an aliquot taken from BD BACTEC Plus and/or BD BACTEC Lytic/10 bottles (Becton Dickinson [BD], Sparks, MD) positive by gram stain for yeast was subcultured to CHROMagar Candida (BD), and a 25-μg fluconazole disk (BD) was placed on the plate. The area of growth inhibition surrounding the disk was measured at 24 and 48 h. In addition, a subculture of the isolate was tested by a microdilution MIC using YeastOne (TREK Diagnostics Systems Inc., OH) and disk diffusion (NCCLS M44-A) using a standardized inoculum plated onto CHROMagar Candida as well as Mueller-Hinton agar to which 2% glucose and 0.5 μg/ml methylene blue dye was added (MH-GMB). The categorical interpretation derived from the MIC was used as the reference to which the disk diffusion results were compared. There were a total of 41
Candida albicans
, 23
Candida glabrata
, 20
Candida parapsilosis
, 9
Candida tropicalis
, and 1 each of
Candida krusei
and
Candida lusitaniae
tested. At 24 h there was full agreement among the methods for all
C. albicans
,
C. tropicalis
,
C. lusitaniae
, and
C. krusei
isolates. For the
C. parapsilosis
isolates at 24 h there was one very major discrepancy using the direct CHROMagar and one major error with the standardized MH-GMB. The majority of the errors were seen at 24 h with the
C. glabrata
isolates. Of the 23
C. glabrata
isolates at 24 h by direct CHROMagar, there were 10 minor and 1 very major error; by MH-GMB there were 12 minor and 2 very major errors; and by standardized CHROMagar Candida there were 13 minor and 2 major errors. There were no very major errors with
C. glabrata
when all plates were read at 48 h. At 24 h by the direct and standardized CHROMagar the majority of
C. glabrata
isolates were more resistant, whereas by MH-GMB they were more susceptible than the reference MIC interpretation. In summary, subculturing yeast directly from blood cultures onto CHROMagar to which a fluconazole disk has been added may provide a presumptive identification at 24 h and, with the exception of
C. glabrata
, was able to predict the susceptibility to fluconazole with the majority of
Candida
isolates examined in this evaluation.
Internally Controlled Real-Time PCR Monitoring of Adenovirus DNA Load in Serum or Plasma of Transplant Recipients
Authors: Claas Eric C. J. , Schilham Marco W. , de Brouwer Caroline S. , Hubacek Petr , Echavarria Marcela , Lankester Arjan C. , van Tol Maarten J. D. , Kroes Aloys C. M. 
Published: April 2005

ABSTRACT

Adenoviruses have been recognized as important pathogens in immunocompromised hosts. Particularly in pediatric allogeneic stem cell transplant recipients, the morbidity of the patients and mortality in those patients with disseminated infections have been found to increase over the last few years. Severe infections are predominantly but not exclusively caused by subgroup C adenoviruses. A multiplex real-time PCR assay using molecular beacons as probes was developed to enable monitoring of adenovirus DNA in those patients with simultaneous identification of subgroups. An internal control was coamplified in the multiplex PCR to check for the DNA isolation procedure as well as the presence of inhibitors in the clinical samples. The assay has been applied retrospectively in patient groups with different clinical outcomes of infection. In fatal cases, significantly higher adenovirus loads developed, exceeding even 10
11
copies/ml of serum or plasma. Patients with viral loads over 10
6
copies/ml appear to have an increased risk for fatal complications. This quantitative real-time PCR assay has been prospectively used clinically since 2002 to study the course of adenovirus infection. In addition, the assay provides objective start and end points of therapeutic interventions, including the clinically important evaluation of antiviral drugs.
Use of an Electronic Nose To DiagnoseMycobacterium bovisInfection in Badgers and Cattle
Authors: Fend R. , Geddes R. , Lesellier S. , Vordermeier H.-M. , Corner L. A. L. , Gormley E. , Costello E. , Hewinson R. G. , Marlin D. J. , Woodman A. C. , Chambers M. A. 
Published: April 2005

ABSTRACTIt is estimated that more than 50 million cattle are infected withMycobacterium bovisworldwide, resulting in severe economic losses. Current diagnosis of tuberculosis (TB) in cattle relies on tuberculin skin testing, and when combined with the slaughter of test-positive animals, it has significantly reduced the incidence of bovine TB. The failure to eradicate bovine TB in Great Britain has been attributed in part to a reservoir of the infection in badgers (Meles meles). Accurate and reliable diagnosis of infection is the cornerstone of TB control. Bacteriological diagnosis has these characteristics, but only with samples collected postmortem. Unlike significant wild animal reservoirs ofM. bovisthat are considered pests in other countries, such as the brushtail possum (Trichosurus vulpecula) in New Zealand, the badger and its sett are protected under United Kingdom legislation (The Protection of Badgers Act 1992). Therefore, an accurate in vitro test for badgers is needed urgently to determine the extent of the reservoir of infection cheaply and without destroying badgers. For cattle, a rapid on-farm test to complement the existing tests (the skin test and gamma interferon assay) would be highly desirable. To this end, we have investigated the potential of an electronic nose (EN) to diagnose infection of cattle or badgers withM. bovis, using a serum sample. Samples were obtained from both experimentally infected badgers and cattle, as well as naturally infected badgers. Without exception, the EN was able to discriminate infected animals from controls as early as 3 weeks after infection withM. bovis, the earliest time point examined postchallenge. The EN approach described here is a straightforward alternative to conventional methods of TB diagnosis, and it offers considerable potential as a sensitive, rapid, and cost-effective means of diagnosingM. bovisinfection in cattle and badgers.
Species of
Phaeoacremonium
Associated with Infections in Humans and Environmental Reservoirs in Infected Woody Plants
Authors: Mostert Lizel , Groenewald Johannes Z. , Summerbell Richard C. , Robert Vincent , Sutton Deanna A. , Padhye Arvind A. , Crous Pedro W. 
Published: April 2005

ABSTRACT

To date, three species of
Phaeoacremonium
have been associated with phaeohyphomycosis. These are
P. parasiticum
(formerly
Phialophora parasitica
),
P. inflatipes
, and
P. rubrigenum
. Numerous unknown isolates resembling
Phaeoacremonium
spp. have in recent years been isolated from human patients as well as from woody plants that appear to be the main environmental source of these fungi. Nine new
Phaeoacremonium
species, of which six were obtained as etiologic agents of human opportunistic infection, are reported. They can be identified based on their cultural and morphological characters, and the identifications are strongly supported in phylogenetic analyses of partial sequences of the actin, β-tubulin, and calmodulin genes. A multiple-entry electronic key based on morphological, cultural, and β-tubulin sequence data was developed to facilitate routine species identification. Reexamination of all isolates of
P. inflatipes
associated with human disease showed them to be misidentified and to belong to the new taxa described here.
Reemergence of
emm
1 and a Changed Superantigen Profile for Group A Streptococci Causing Invasive Infections: Results from a Nationwide Study
Authors: Ekelund Kim , Skinhøj Peter , Madsen Jesper , Konradsen Helle Bossen 
Published: April 2005

ABSTRACT

Between 1999 and 2002, 496 invasive group A streptococcal (GAS) isolates from clinical microbiological departments in Denmark and subsequently 487 (98%) questionnaires from the clinicians treating the patients were received as part of a national surveillance.
emm
types and streptococcal superantigen (SAg) genes were determined. The incidence of invasive GAS infections was on average 2.3 per 100,000 per year. Bacteremia with no focal symptoms (27%) was together with erysipelas (20%) the most prevalent clinical diagnoses. Streptococcal toxic shock syndrome occurred in 10% of patients, of which 56% died. The overall case fatality rate within 30 days was 23%. In total, 47 different
emm
types were identified, of which
emm
1,
emm
3,
emm
4,
emm
12,
emm
28, and
emm
89 were identified in 72% of the 493 available isolates. During the 4-year period the presence of
emm
1 increased from 16% in 1999 to 40% in 2002. Concurrently, the presence of
emm
3 decreased from 23% in 1999 to 2% in 2002. The
emm
1 isolates predominantly carried
speA
, although the frequency decreased from 94% in 1999 to 71% in 2002, whereas the
emm
1-specific prevalence of
speC
increased from 25 to 53%. In a historical perspective, this could be interpreted as a reemergence of
emm
1 and could indicate a possible introduction of a new
emm
1 subclone. However, this reemergence did not result in any significant changes in the clinical manifestations during the study period. Our results show the complexity of invasive GAS infections, with time-dependent variations in the incidence and distribution of
emm
and SAg genes, which emphasizes the need for continuous epidemiological and molecular investigations.