bluuebunny/crossref_metadata_2025_split huggingface dataset:
Prediction of Decreased Susceptibility to Penicillin of
Neisseria meningitidis
Strains by Real-Time PCR
Authors: Stefanelli Paola , Carattoli Alessandra , Neri Arianna , Fazio Cecilia , Mastrantonio Paola 
Published: October 2003

ABSTRACT

Sequence analysis of the
penA
gene, encoding penicillin-binding protein 2 (PBP2), in 30 penicillin-intermediate (PenI)
Neisseria meningitidis
strains showed altered gene sequences due to the translocation of exogenous DNA blocks derived from commensal neisseriae, which are known to have PBP2 proteins with decreased affinity for the antibiotic. In order to obtain a rapid and reproducible method for predicting the PenI phenotype, a real-time PCR assay was set up with primers and probes designed on the basis of the
penA
gene. The A→G mutation at codon 566, in the transpeptidase domain of the
penA
gene (which is present in the whole sample of 30 PenI strains and in all the 41 sequences of PenI meningococci isolated worldwide and has been deposited in the sequence databank), was chosen as a marker of
penA
translocations. Two hybridization probes were designed to distinguish the wild-type
penA
gene in penicillin-susceptible (PenS) meningococci from the mutated
penA
gene at codon 566 in PenI strains. Thermal analysis of probe hybridization revealed a melting temperature difference of at least 6°C between PenI and PenS strains. This real-time PCR protocol characterizes the penicillin phenotype of
N. meningitidis
in a few hours without DNA sequencing and is useful for rapid screening of the penicillin-intermediate genotype among meningococcal isolates.
Comparison of a Shiga Toxin Enzyme-Linked Immunosorbent Assay and Two Types of PCR for Detection of Shiga Toxin-Producing
Escherichia coli
in Human Stool Specimens
Authors: Pulz Matthias , Matussek Andreas , Monazahian Masyar , Tittel Andreas , Nikolic Elisabet , Hartmann Maike , Bellin Tobias , Buer Jan , Gunzer Florian 
Published: October 2003

ABSTRACT

Shiga toxin (Stx)-producing
Escherichia coli
(STEC) is a major cause of sporadic cases of disease as well as serious outbreaks worldwide. The spectrum of illnesses includes mild nonbloody diarrhea, hemorrhagic colitis, and hemolytic-uremic syndrome. STEC produces one or more Stxs, which are subdivided into two major classes, Stx1 and Stx2. The ingestion of contaminated food or water, person-to-person spread, and contact with animals are the major transmission modes. The infective dose of STEC may be less than 100 organisms. Effective prevention of infection is dependent on rapid detection of the causative bacterial pathogen. In the present study, we examined 295 stool specimens for the presence of Stx-producing
E
.
coli
by three different methods: an Stx enzyme-linked immunosorbent assay, a conventional PCR assay, and a LightCycler PCR (LC-PCR) assay protocol recently developed by our laboratory at the Institute of Medical Microbiology at Hannover Medical School. Our intent was to compare these three methods and to examine the utility of the STEC LC-PCR protocol in a clinical laboratory. The addition of a control DNA to each sample to clearly discriminate inhibited specimens from negative ones enhanced the accuracy of the LC-PCR protocol. From our results, it can be concluded that LC-PCR is a very useful tool for the rapid and safe detection of STEC in clinical samples.
Genotypic Analyses of
Vibrio parahaemolyticus
and Development of a Pandemic Group-Specific Multiplex PCR Assay
Authors: Okura Masatoshi , Osawa Ro , Iguchi Atsushi , Arakawa Eiji , Terajima Jun , Watanabe Haruo 
Published: October 2003

ABSTRACT

A total of 54
Vibrio parahaemolyticus
strains including pandemic O3:K6 strains and newly emerged O4:K68, O1:K25, O1:K26, and O1:K untypeable strains (collectively referred to as the “pandemic group”) were examined for their pulsed-field gel electrophoresis (PFGE) and arbitrarily primed PCR (AP-PCR) profiles and for the presence or absence of genetic marker DNA sequences,
toxRS
/
new
or
orf8
, that had been reported elsewhere to be specific for the pandemic group. Both PFGE and AP-PCR analyses indicated that all strains of the pandemic group formed a distinct genotypic cluster, suggesting that they originated from the same clone. In addition to the pandemic group, four O3:K6 strains that did not possess the thermostable direct hemolysin (
tdh
) gene also belonged to this cluster and possessed the
toxRS
/
new
sequence. However, three O3:K6 strains that clearly belonged to the pandemic group by PFGE and AP-PCR did not possess the
orf8
sequence. The evidence suggests that neither the
toxRS
/
new
nor the
orf8
sequence is a reliable gene marker for definite identification of the pandemic group. We therefore developed a novel multiplex PCR assay specific for the pandemic group. The assay successfully distinguished pandemic group strains from other
V. parahaemolyticus
strains by yielding two distinct PCR products for
tdh
(263 bp) and the
toxRS
/
new
sequence (651 bp).
Description of a New Yeast Species,
Malassezia japonica
, and Its Detection in Patients with Atopic Dermatitis and Healthy Subjects
Authors: Sugita Takashi , Takashima Masako , Kodama Minako , Tsuboi Ryoji , Nishikawa Akemi 
Published: October 2003

ABSTRACT

Lipophilic yeasts of the genus
Malassezia
are part of the normal cutaneous microflora and are considered one of the factors that trigger atopic dermatitis (AD). We isolated two strains of
Malassezia
from a healthy Japanese female. Analysis of the D1/D2 26S ribosomal DNA and internal transcribed spacer region sequences of the isolates suggested that they are new members of the genus
Malassezia
. We propose the name
Malassezia japonica
sp. nov. for the isolates.
M
.
japonica
is easily distinguished from the seven known lipophilic species by its ability to assimilate Tween 40 and Tween 60 and its inability to assimilate Tween 20 and Tween 80 and to grow at 40°C. Furthermore, by applying transparent dressings to the skin lesions of 36 patients with AD and the skin of 22 healthy subjects,
M
.
japonica
DNA was detected by a non-culture-based method consisting of nested PCR with
M
.
japonica
species-specific primers.
M
.
japonica
DNA was detected from 12 of the 36 patients (33.3%) and 3 of the 22 healthy subjects (13.6%). Although it is not known whether
M
.
japonica
plays a role in AD, this species was part of the microflora in both patients with AD and healthy subjects.
Diagnosis of
Helicobacter pylori
Infection in Adults with Intellectual Disability
Authors: Wallace Robyn A. , Schluter Philip J. , Forgan-Smith Ross , Wood Robyn , Webb Penelope M. 
Published: October 2003

ABSTRACT

Helicobacter pylori
infection is common among adults with intellectual disability. The acceptabilities and accuracies of different diagnostic tests in this population are unknown. We aimed to determine (i) patient acceptability and (ii) performance characteristics of serology, fecal-antigen, and urea breath tests among adults with intellectual disability. One hundred sixty-eight such adults underwent
H. pylori
testing with serology and fecal-antigen tests, and a portion underwent treatment. One year later, the participants were retested with fecal-antigen, serology, and urea breath tests. The numbers of specimens obtained and difficulties in collection reported by caregivers were noted. Test performance characteristics were assessed among participants and 65 of their caregivers, using serology as the reference. All participants provided at least one specimen, despite reported collection difficulties for 23% of fecal and 27% of blood specimens. Only 25% of the participants provided breath specimens; failure to perform this test was associated with lower intellectual ability and higher maladaptive behavior. The sensitivity, specificity, and positive and negative predictive values of the fecal test (baseline and 12 months versus caregivers) were 70 and 63 versus 81, 93 and 95 versus 98, 96 and 92 versus 93, and 53 and 74 versus 93%, respectively; those of the urea breath test (12 months versus caregivers) were 86 versus 100, 88 versus 95, 75 versus 89, and 94 versus 100%, respectively. With assistance, fecal or blood specimens for
H. pylori
assessment can be provided by most patients with intellectual disability regardless of their level of function or behavior. Only those with greater ability can perform the urea breath test. Using serology as the reference test, the limitations of performance characteristics of the fecal-antigen and urea breath tests are similar to those among a control group of caregivers.
Evaluation of the VITEK 2 System for Rapid Direct Identification and Susceptibility Testing of Gram-Negative Bacilli from Positive Blood Cultures
Authors: Ling Thomas K. W. , Liu Z. K. , Cheng Augustine F. B. 
Published: October 2003

ABSTRACT
This study explores the possibility of combining the BacT/Alert Microbial Detection System with the VITEK 2 system to achieve rapid bacterial identification and susceptibility testing. Direct inoculation of bacterial suspension to the VITEK 2 ID-GNB card and AST-NO09 card was made by differential centrifugation of blood cultures of organisms with gram-negative enteric bacillus-like morphology. A total of 118 strains were investigated; of these, 97 (82.2%) strains were correctly identified to the species level and 21 (17.8%) strains were not identified; by comparing the results with those of the reference method of API identification systems using a pure culture, it was found that no strain had been misidentified. Among the 21 strains with no identification, 13 (61.9%) strains were nonfermenters. The direct-identification reporting time of VITEK 2 was 3.3 h. Direct testing of susceptibility to 11 antibiotics, i.e., amikacin, cefepime, ceftazidime, ciprofloxacin, gentamicin, imipenem, meropenem, netilmicin, piperacillin, piperacillin-tazobactam, and tobramycin, was also performed by using the broth microdilution (MB) method according to the NCCLS guidelines as a reference. After comparing the MICs of the VITEK 2 system with those obtained by the MB method within ±twofold dilution, it was determined that the 1,067 organism-antibiotic combinations had an overall correct rate of 97.6% (1,041 combinations). The rates of susceptibility to the 11 antibiotics ranged from 88.7 to 100%, respectively. Only two (0.2%) and four (0.4%) combinations of the susceptibility tests gave very major errors (i.e., reported as sensitive by the VITEK 2 system but shown to be resistant by the MB method) and major errors (i.e., reported as resistant by the VITEK 2 system but shown to be sensitive by the MB method), respectively. The reporting time for the direct testing of susceptibility against the 11 antibiotics for 97 blood culture isolates by the VITEK 2 system ranged from 3.3 to 17.5 h. Compared with conventional methods that require 1 or 2 days, this method can make same-day reporting possible and thus permit better patient management.
Algorithm To Determine Cost Savings of Targeting Antimicrobial Therapy Based on Results of Rapid Diagnostic Testing
Authors: Oosterheert J. J. , Bonten M. J. M. , Buskens E. , Schneider M. M. E. , Hoepelman I. M. 
Published: October 2003

ABSTRACT

A rapid diagnosis of pneumococcal pneumonia may allow the earlier use of narrow-spectrum antimicrobial therapy. It is unknown, however, whether rapid diagnostic testing of patients hospitalized with community-acquired pneumonia (CAP) admitted to hospital lowers costs. Therefore, an algorithm to calculate the costs associated with the diagnosis and treatment of CAP was formulated. Subsequently, the algorithm was applied to clinical data for 122 consecutively admitted patients with CAP whose sputum samples were Gram stained and whose urine was tested for
Streptococcus pneumoniae
antigen. The costs of initial antimicrobial therapy, personnel, and materials were measured. Compared to the most expensive empirical regimen, rapid diagnostic testing would result in cost savings per patient (PP) of €3.51 for Gram staining and €8.11 for urinary pneumococcal antigen testing (€1 is equal to US$1.13, from 2000 to 2002). Compared to the cheapest regimen, Gram staining would increase the cost by €2.25 PP, and urinary antigen testing would increase the cost by €24.26 PP. In our setting, the use of rapid diagnostic testing would not lower costs. Cost savings depend, however, on the differences in the prices of the different antibiotics chosen and the proportion of evaluable and positive samples.
Detection of Simulated Candidemia by the BACTEC 9240 System with Plus Aerobic/F and Anaerobic/F Blood Culture Bottles
Authors: Horvath Lynn L. , Hospenthal Duane R. , Murray Clinton K. , Dooley David P. 
Published: October 2003

ABSTRACT

We studied the ability of the BACTEC 9240 automated blood culture system to detect simulated candidemia, including both
Candida albicans
and non-
albicans Candida
species. Simulated blood cultures were produced using 50
Candida
isolates and BACTEC Plus Aerobic/F and Anaerobic/F blood culture bottles. Ten milliliters of blood and a suspension of each isolate containing 1,000 CFU were introduced into each bottle and then incubated at 35°C in the BACTEC 9240 system. The system detected growth in 56 of 100 bottles. Four isolates did not have growth detected in either bottle after 21 days of incubation, resulting in four missed episodes of candidemia. If the blood culture bottles had been incubated for 5 days, an additional episode of candidemia would have remained undetected. If the bottles had been incubated for only 3 days, another episode would have been missed, resulting in up to six missed episodes of candidemia (four
Candida glabrata
isolates, one
C. albicans
isolate, and one
Candida rugosa
isolate). Terminal subculture of bottles without detected growth recovered yeast in 93% (41 of 44) of the bottles, representing 41 false negatives. In bottles where growth was detected, the time to detection was ∼24 h. However, the mean time to growth detection for
C. glabrata
isolates in anaerobic medium was 22.14 ± 2.47 h, but it was 120.89 ± 35.33 h in aerobic medium (
P
< 0.001). The BACTEC 9240 system detected growth of most
Candida
isolates; however, the delayed time to detection of
C. glabrata
is clinically significant. Given the high rate of false negatives, terminal subcultures may be helpful in certain situations.
Extended Epidemic of Nosocomial Urinary Tract Infections Caused by
Serratia marcescens
Authors: Su Lin-Hui , Ou Jonathan T. , Leu Hsieh-Shong , Chiang Ping-Cherng , Chiu Yueh-Pi , Chia Ju-Hsin , Kuo An-Jing , Chiu Cheng-Hsun , Chu Chishih , Wu Tsu-Lan , Sun Chien-Feng , Riley Thomas V. , Chang Barbara J. 
Published: October 2003

ABSTRACT

In recent years a significant increase in the incidence of
Serratia marcescens
infections was noted at the Chang Gung Memorial Hospital, Taoyuan, Taiwan. A review of laboratory (1991 to 2002) and infection control (1995 to 2002) records showed the possibility of an extended epidemic of nosocomial urinary tract infections (UTIs) caused by
S. marcescens
. Therefore, in 1998 and 1999, 87 isolates were collected from patients with such infections and examined and another 51 isolates were collected in 2001 and 2002. The patients were mostly elderly or the infections were associated with the use of several invasive devices.
S. marcescens
was usually the only pathogen found in urine cultures in our study. Neither prior infections nor disseminated infections with the organism were observed in these patients. Resistance to most antibiotics except imipenem was noted. Two genotyping methods, pulsed-field gel electrophoresis and infrequent-restriction-site PCR, were used to examine the isolates. A total of 12 genotypes were identified, and 2 predominant genotypes were found in 72 (82.8%) of the 87 isolates derived from all over the hospital. However, 63.9% of the isolates of the two genotypes were from neurology wards. A subsequent intervention by infection control personnel reduced the infection rate greatly. The number and proportion of the two predominant genotypes were significantly reduced among the 51 isolates collected in 2001 and 2002. Thus, a chronic and long-lasting epidemic of nosocomial UTIs caused by
S. marcescens
was identified and a successful intervention was carried out. Both a cautious review of laboratory and infection control data and an efficient genotyping system are necessary to identify such a cryptic epidemic and further contribute to the quality of patient care.
Utility of Multilocus Sequence Typing as an Epidemiological Tool for Investigation of Outbreaks of Gastroenteritis Caused by
Campylobacter jejuni
Authors: Sails Andrew D. , Swaminathan Bala , Fields Patricia I. 
Published: October 2003

ABSTRACT

Multilocus sequence typing (MLST) has been proven useful for the study of the global population structure of
Campylobacter jejuni
; however, its usefulness for the investigation of outbreaks of disease caused by
C. jejuni
has not been proven. In this study, MLST plus sequencing of the
flaA
short variable region (SVR) were applied to 47 isolates from 12 outbreaks of
C. jejuni
infection whose relatedness has been determined previously, and the results were compared to those of serotyping and pulsed-field gel electrophoresis (PFGE). Isolates implicated in an outbreak were indistinguishable by all four subtyping methods, with sporadic isolates being distinguished from outbreak isolates. Two sporadic isolates from one outbreak were resistant to
Sma
I digestion and therefore nontypeable by PFGE but were differentiated from the outbreak strain by the other methods. PFGE and
flaA
SVR typing were the most discriminatory methods, with discriminatory indices (DI) of 0.930 and 0.923, respectively. However, an epidemic strain from one outbreak was distinguished from the other outbreak isolates by
flaA
SVR typing; its
flaA
allele was different at five nucleotides, suggesting that this change was possibly mediated by recombination. MLST was less discriminatory than PFGE and
flaA
SVR typing (DI = 0.859), and many of the epidemic strains possessed common sequence types (STs) including ST-8, -21, -22, and -42. However, further discrimination within STs was achieved by
flaA
SVR typing or PFGE. The results from this study demonstrate that a combined approach of MLST plus
flaA
SVR typing provides a level of discrimination equivalent to PFGE for outbreak investigations.
Practical Disk Diffusion Method for Detection of Inducible Clindamycin Resistance in
Staphylococcus aureus
and Coagulase-Negative Staphylococci
Authors: Fiebelkorn K. R. , Crawford S. A. , McElmeel M. L. , Jorgensen J. H. 
Published: October 2003

ABSTRACT

Resistance to macrolides in staphylococci may be due to active efflux (encoded by
msrA
) or ribosomal target modification (macrolide-lincosamide-streptogramin B [MLS
B
] resistance; usually encoded by
ermA
or
ermC
). MLS
B
resistance is either constitutive or inducible following exposure to a macrolide. Induction tests utilize closely approximated erythromycin and clindamycin disks; the flattening of the clindamycin zone adjacent to the erythromycin disk indicates inducible MLS
B
resistance. The present study reassessed the reliability of placing erythromycin and clindamycin disks in adjacent positions (26 to 28 mm apart) in a standard disk dispenser, compared to distances of 15 or 20 mm. A group of 130 clinical isolates of
Staphylococcus aureus
and 100 isolates of erythromycin-resistant coagulase-negative staphylococci (CNS) were examined by disk approximation; all CNS isolates and a subset of
S. aureus
isolates were examined by PCR for
ermA
,
ermC
, and
msrA
. Of 114 erythromycin-resistant
S. aureus
isolates, 39 demonstrated constitutive resistance to clindamycin, while 33 showed inducible resistance by disk approximation at all three distances. Only one isolate failed to clearly demonstrate induction at 26 mm. Of 82 erythromycin-resistant CNS isolates that contained
ermA
or
ermC
, 57 demonstrated constitutive clindamycin resistance, and 25 demonstrated inducible resistance, at 20 and 26 mm. None of the 42
S. aureus
isolates or 18 CNS isolates containing only
msrA
and none of the erythromycin-susceptible isolates yielded positive disk approximation tests. Simple placement of erythromycin and clindamycin disks at a distance achieved with a standard disk dispenser allowed detection of 97% of
S. aureus
strains and 100% of CNS strains with inducible MLS
B
resistance in this study.
Direct Susceptibility Testing of Positive Blood Cultures by Using Sensititre Broth Microdilution Plates
Authors: Chapin Kimberle C. , Musgnug Michael C. 
Published: October 2003

ABSTRACT
Traditional susceptibility testing of blood cultures requires overnight incubation in order to obtain isolated colonies. Susceptibility results can be reported up to 24 h sooner by using a bacterial pellet from the blood culture broth. This study evaluated the accuracy of direct susceptibility testing from positive ESP blood culture broths by using Sensititre broth microdilution plates compared to testing with isolated colonies. Practical inclusion criteria were applied to gram-positive organisms to avoid reporting susceptibilities for probable contaminants. All gram-negative organisms were tested directly. An aliquot of the blood culture was centrifuged, and the resulting pellet was used to make a 0.5 McFarland suspension. Microdilution plates were inoculated and interpreted according to the manufacturer's instructions. Colony counts were performed to ensure proper colony density was achieved. A total of 199 patient and seeded blood cultures were evaluated for both essential (within ±1 twofold dilution) and categorical (sensitive, intermediate, or resistant) agreement. Testing of 93 gram-positive isolates (1,214 antimicrobial agent-organism combinations) yielded 98% essential agreement and categorical error rates of 0.3% minor, no major (false resistance), and 1.7% very major (false susceptibility) errors. For 106 gram-negative isolates (1,828 antimicrobial agent-organism combinations), the essential agreement was 99%. Categorical error rates were 0.5, 0, and 2.0% for minor, major, and very major errors, respectively. Performance was comparable for both gram-positive and gram-negative isolates, as well as for both aerobic and anaerobic media. Using this direct testing methodology, reliable susceptibility results can be reported to physicians 24 h sooner, allowing earlier appropriate modification of antimicrobial therapy.
Microwave Sterilization of Femoral Head Allograft
Authors: Dunsmuir Robert A. , Gallacher Grace 
Published: October 2003

ABSTRACTThe potential shortage of allograft bone has led to the need to investigate other sources of bone for allografts. Some allograft bone donated from primary total hip arthroplasty recipients must be discarded or treated to become useable as a result of bacterial contamination. Femoral head allografts were contaminated withStaphylococcus aureusandBacillus subtilis. A domestic microwave oven was used. The contaminated bone was exposed to microwave irradiation for different time periods. The samples were then cultured to attempt to grow the two bacterial species. The contaminated bone samples failed to grow any organisms after 2 min of exposure to microwave irradiation. This study shows that sterilization of femoral head allografts contaminated withS. aureusandB. subtiliscan be achieved with microwave irradiation in a domestic microwave oven. This method of sterilization of bone allografts is cheap, easily used, and an effective way to process contaminated bone.