bluuebunny/crossref_metadata_2025_split huggingface dataset:
Lack of Utility of the Lysis-Centrifugation Blood Culture Method for Detection of Fungemia in Immunocompromised Cancer Patients
Authors: Creger Richard J. , Weeman Kisa E. , Jacobs Michael R. , Morrissey Anne , Parker Pamela , Fox Robert M. , Lazarus Hillard M. 
Published: December 2019

ABSTRACT
We retrospectively compared the utility of a fungal isolation device (Isolator) versus conventional techniques for recovering fungal organisms from blood cultures obtained from neutropenic cancer patients. Positive cultures were deemed true pathogens, possible pathogens, or contaminants according to laboratory and clinical criteria. Fifty-three patients had 66 positive blood cultures for fungi, nine on multiple occasions. In 20 episodes true pathogens were recovered, 6 from broth medium alone, 4 from the Isolator system alone, and 10 from both systems. False-negative cultures were noted in 4 of 20 (20%) cases in which broth medium was used and in 6 of 20 (30%) cases in which the Isolator system was used. Possible pathogens were detected in 4 of 66 blood culture-positive cases. Forty-two positive cultures were considered contaminants, 1 collected from standard medium and 41 of 42 (98%) which grew only in Isolators. Eleven of 18 patients with true fungal infections expired as a result of infection, while 4 of 33 patients with a contaminant expired, none from a fungal cause. We do not advocate the routine use of Isolator tubes in the evaluation of the febrile, neutropenic patient due to the high rates of false positives and of contamination.
Detection of Measles Virus mRNA from Autopsied Human Tissues
Authors: Katayama Yuko , Kohso Kiyoyasu , Nishimura Akiyoshi , Tatsuno Yoshitsugu , Homma Morio , Hotta Hak 
Published: December 2019

ABSTRACT
By reverse transcription-PCR, measles virus (MV) mRNA was detected in the brain, kidney, spleen, liver, and lung tissues obtained from 23 (45.1%) of 51 autopsy subjects, with the detection rates of each tissue ranging from 8 to 20%. Sequence analysis revealed frequent mutations in the corresponding viral protein. These results suggest that MV mutants commonly persist in apparently healthy individuals.
Identification of
Encephalitozoon intestinalis
in Travelers with Chronic Diarrhea by Specific PCR Amplification
Authors: Raynaud Laurent , Delbac Frederic , Broussolle Veronique , Rabodonirina Meja , Girault Veronique , Wallon Martine , Cozon Gregoire , Vivares Christian P. , Peyron François 
Published: December 2019

ABSTRACT

With the use of Weber’s modified trichrome and Uvitex 2B techniques, spores of microsporidia were detected in the stools of four travelers presenting clinically with chronic diarrhea. The general health of these patients was not impaired, and human immunodeficiency virus screening was negative. Immune evaluation, including the study of lymphocytic subpopulations, assay of serum immunoglobulins, and an intradermal multitest, showed normal results. Molecular identification of microsporidian species was based on the PCR amplification of a small-subunit rRNA sequence followed by
Hin
fI endonuclease restriction.
Encephalitozoon intestinalis
microsporidiosis was thus shown in two of the four patients examined. In two patients, therapy based on albendazole made stools devoid of microsporidian spores without influence on the intestinal disorders. The pathogenic role of
E. intestinalis
in immunocompetent individuals remains to be demonstrated.
Characterization of Gentamicin-Susceptible Strains of Methicillin-Resistant
Staphylococcus aureus
Involved in Nosocomial Spread
Authors: Lemaître Nadine , Sougakoff Wladimir , Masmoudi Afef , Fievet Marie-Hélène , Bismuth Roland , Jarlier Vincent 
Published: December 2019

ABSTRACT

We report an outbreak of epidemic
Staphylococcus aureus
strains characterized by an unusual heterogeneous resistance to methicillin and resistance to tobramycin but susceptibility to gentamicin (gentamicin-susceptible methicillin-resistant
S. aureus
[GS-MRSA]), contrasting with gentamicin-resistant homogeneous MRSA (GR-MRSA) that have been endemic in our hospital since the 1970s. A total of 97 GS-MRSA strains, which were shown by DNA hybridization to carry the
mecA
and
ant(4′)-Ia
genes, were studied. The 40 GS-MRSA strains isolated at the beginning of the outbreak (January 1992 to June 1993) were typed by using resistance patterns, phage typing, serotyping, and pulsed-field gel electrophoresis and were compared with GR-MRSA and methicillin-susceptible
S. aureus
(MSSA) strains isolated during the same period. Two dominant clones, A::1 and B::3, and one minor clone, C::5, were identified among the 40 GS-MRSA strains, according to pulsotypes (A to C) and their resistance patterns (1, 3, and 5), which were distinguishable from those of GR-MRSA and MSSA strains. A selection of 57 GS-MRSA strains, isolated from 1994 to 1996, were clustered in the same three clones. However, their distribution had changed in comparison with that in the 1992 to 1993 period: clone A::1 remained dominant (47 versus 42.5%), whereas clone B::3 progressively declined (5 versus 35%) and clone C::5, the most susceptible to antibiotics, spread (44 versus 2.5%). Epidemiological investigations revealed that some clones had been introduced via patients transferred from other hospitals and that cross-infection occurred within and between wards. Major changes in the use of antibiotics, especially aminoglycosides, cyclines, and macrolides, likely played a role in the emergence and spread of GS-MRSA strains.
Use of an Enrichment Broth Cultivation-PCR Combination Assay for Rapid Diagnosis of Swine Erysipelas
Authors: Shimoji Yoshihiro , Mori Yasuyuki , Hyakutake Koji , Sekizaki Tsutomu , Yokomizo Yuichi 
Published: December 2019

ABSTRACT

We have previously described the creation by Tn
916
mutagenesis of avirulent transposition mutants from a highly virulent strain of
Erysipelothrix rhusiopathiae
, the causative agent of swine erysipelas. In this study, we cloned a 2.2-kb DNA fragment which flanked the Tn
916
insertion in an avirulent mutant (strain 33H6) and evaluated the possibility that this region could be used for the specific detection of
E. rhusiopathiae
. According to the sequences of this region, oligonucleotide primers were designed to amplify a 937-bp fragment of the
E. rhusiopathiae
chromosome by PCR. The specificity of the PCR was investigated by analyzing 64 strains of
Erysipelothrix
species and 27 strains of other genera different from
Erysipelothrix
. A 937-bp DNA fragment could be amplified from all
E. rhusiopathiae
strains tested, and no amplification was observed by using DNAs from the other species tested. To make a rapid and definite diagnosis of swine erysipelas in slaughterhouses, we developed an enrichment broth cultivation-PCR combination assay, which used a commercially available DNA extraction kit, to identify
E. rhusiopathiae
in the specimens from swine with arthritis. After samples were enriched in selective broth culture, detection of
E. rhusiopathiae
was tested by either conventional methods or the PCR. Of 102 samples tested, 15 samples were positive by conventional methods and 12 of the 15 samples were positive by the PCR. The detection limit of the PCR was 10
3
CFU per reaction mixture for the PCR-positive samples. These results indicate that this PCR technique could be used as a first-line screening technique for the specific detection of
E. rhusiopathiae
in specimens.
Detection of Resistance to Amphotericin B among
Cryptococcus neoformans
Clinical Isolates: Performances of Three Different Media Assessed by Using E-Test and National Committee for Clinical Laboratory Standards M27-A Methodologies
Authors: Lozano-Chiu M. , Paetznick V. L. , Ghannoum M. A. , Rex J. H. 
Published: December 2019

ABSTRACT

Although reliable detection of resistance in vitro is critical to the overall performance of any susceptibility testing method, the recently released National Committee for Clinical Laboratory Standards M27-A methodology for susceptibility testing of yeasts discriminates poorly between resistant and susceptible isolates of
Candida
spp. We have previously shown that both substitution of antibiotic medium 3 for RPMI 1640 medium in the microdilution variant of the M27-A method and use of the E-test agar diffusion methodology permit detection of amphotericin B-resistant
Candida
isolates. To determine the relevance of these observations to
Cryptococcus neoformans
, we have evaluated the performances of both the M27-A and the E-test methodologies with this yeast using three different media (RPMI 1640 medium, antibiotic medium 3, and yeast nitrogen base). As with
Candida
, we found that only antibiotic medium 3 permitted consistent detection of resistant isolates when testing was performed in broth by the M27-A method. When testing was performed by the E-test agar diffusion method, both RPMI 1640 medium and antibiotic medium 3 agar permitted ready detection of the resistant isolates. Reading of the results after 48 h of incubation was required for testing in broth by the M27-A method, while the MIC could be determined after either 48 or 72 h when the agar diffusion method was used.
Rapid Shell Vial Culture Technique for Detection of Enteroviruses and Adenoviruses in Fecal Specimens: Comparison with Conventional Virus Isolation Method
Authors: Van Doornum G. J. J. , De Jong J. C. 
Published: December 2019

ABSTRACT
Detection of enteroviruses and adenoviruses mainly in fecal specimens by rapid culture with inoculation onto cell monolayers in flat-bottom tubes by centrifugation and immunofluorescence staining with genus-specific monoclonal antibodies was compared with that by the conventional virus isolation procedure. For both conventional culture and shell vial culture human lung fibroblast cells and tertiary monkey kidney cells were used. For enterovirus detection, 979 clinical specimens (916 stool specimens, 56 cerebrospinal fluid specimens, and 7 nasopharyngeal swabs) were used. Conventional culture detected 74 enterovirus isolates. A cytopathic effect compatible with the presence of an enterovirus after 3 days of incubation occurred in 25 of the 74 (34%) specimens that eventually became positive. The detection rate for enteroviruses by rapid cell culture after 2 to 3 days of incubation was 42 of 74 (57%). The genus-specific enterovirus monoclonal antibody did not react with strains of echovirus types 22 and 23 or enterovirus type 71. Rapid cell culture for the detection of adenoviruses was performed with 567 clinical specimens (536 stool specimens, 25 cerebrospinal fluid specimens, and 6 miscellaneous specimens), in which 42 adenoviruses were found by conventional culture. Nine of the 42 (21%) adenovirus isolates were detected by conventional culture within 3 days after inoculation, whereas 21 (50%) were found by rapid cell culture within 2 to 3 days. Only two of the nine specimens found to be positive for the enteric adenovirus type 41 by conventional culture as well by a type-specific enzyme-linked immunosorbent assay (ELISA) tested positive by rapid cell culture. In conclusion, the rapid shell vial assay allows the early detection and identification of enteroviruses and adenoviruses in clinical specimens but is markedly less sensitive than the conventional isolation procedure according to the eventual results of the conventional isolation procedure. Conventional cell culture remains a prerequisite for serotyping of enteroviral isolates. On the basis of the results for adenovirus type 41, the rapid detection of adenoviruses was not considered to be useful for the detection of clinically relevant adenoviruses in fecal samples.
Direct Identification ofVibrio vulnificusin Clinical Specimens by Nested PCR
Authors: Lee Shee Eun , Kim Soo Young , Kim Sei Jong , Kim Hyun Soo , Shin Jong Hee , Choi Sang Ho , Chung Sun Sik , Rhee Joon Haeng 
Published: December 2019

ABSTRACTThis study was performed to establish optimal nested PCR conditions and a high-yield DNA extraction method for the direct identification ofVibrio vulnificusin clinical specimens. We designed two sets of primers targeting theV. vulnificushemolysin/cytolysin gene. The target of the first primer set (P1-P2; sense, 5′-GAC-TAT-CGC-ATC-AAC-AAC-CG-3′, and antisense, 5′-AGG-TAG-CGA-GTA-TTA-CTG-CC-3′, respectively) is a 704-bp DNA fragment. The second set (P3-P4; sense, 5′-GCT-ATT-TCA-CCG-CCG-CTC-AC-3′, and antisense, 5′-CCG-CAG-AGC-CGT-AAA-CCG-AA-3′, respectively) amplifies an internal 222-bp DNA fragment. We developed a direct DNA extraction method that involved boiling the specimen pellet in a 1 mM EDTA–0.5% Triton X-100 solution. The new DNA extraction method was more sensitive and reproducible than other conventional methods. The DNA extraction method guaranteed sensitivity as well, even whenV. vulnificuscells were mixed with other bacteria such asEscherichia coliorStaphylococcus aureus. The nested PCR method could detect as little as 1 fg of chromosomal DNA and single CFU ofV. vulnificus. We applied the nested PCR protocol to a total of 39 serum specimens and bulla aspirates from septicemic patients. Seventeen (94.4%) of the 18V. vulnificusculture-positive specimens were positive by the nested PCR. Eight (42.1%) of the 19 culture-negative samples gave positive nested PCR results.
Outbreak of
Pseudomonas fluorescens
Bacteremia among Oncology Patients
Authors: Hsueh Po-Ren , Teng Lee-Jene , Pan Hui-Ju , Chen Yu-Chi , Sun Chun-Chuan , Ho Shen-Wu , Luh Kwen-Tay 
Published: December 2019

ABSTRACT

From 7 to 24 March 1997, four patients developed
Pseudomonas fluorescens
bacteremia at the hospital; one on the oncology ward and the other three in the chemotherapy room. These patients all had underlying malignancies and had the Port-A-Cath (Smiths Industries Medical Systems, Deltec, Inc., St. Paul, Minn.) implants. Three patients had primary bacteremia, and one had Port-A-Cath-related infection. None of these patients had received a blood transfusion before the episodes of bacteremia. All patients recovered: two received antimicrobial agents with in vitro activity against the isolates, and the other two did not have any antibiotic treatment. A total of eight blood isolates were recovered from these patients during the febrile episodes that occurred several minutes after the infusion of chemotherapeutic agents via the Port-A-Cath. These isolates were initially identified as
P. fluorescens
or
Pseudomonas putida
(four),
Burkholderia
(
Ralstonia
)
pickettii
(three), and a non-glucose-fermenting gram-negative bacillus (one) by routine biochemical methods and the Vitek GNI card. These isolates were later identified as
P. fluorescens
on the basis of the characteristic cellular fatty acid chromatogram and the results of supplemental biochemical tests. The identification of identical antibiotypes by the E test and the random amplified polymorphic DNA patterns generated by arbitrarily primed PCR of the isolates showed that the outbreak was caused by a single clone of
P. fluorescens
. Surveillance cultures of the possibly contaminated infusion fluids and disinfectants, which were performed 7 days after recognition of the last infected patient, failed to isolate
P. fluorescens
. This report of a small outbreak caused by
P. fluorescens
suggests that timely, accurate identification of unusual nosocomial pathogens is crucial for early initiation of an epidemiological investigation and timely control of an outbreak.
Molecular Genetic Variation in
Emmonsia crescens
and
Emmonsia parva
, Etiologic Agents of Adiaspiromycosis, and Their Phylogenetic Relationship to
Blastomyces dermatitidis
(
Ajellomyces dermatitidis
) and Other Systemic Fungal Pathogens
Authors: Peterson Stephen W. , Sigler Lynne 
Published: December 2019

ABSTRACT

Emmonsia crescens
, an agent of adiaspiromycosis,
Blastomyces dermatitidis
, the agent of blastomycosis, and
Histoplasma capsulatum
, the agent of histoplasmosis, are known to form meiotic (sexual) stages in the ascomycete genus
Ajellomyces
(
Onygenaceae
,
Onygenales
), but no sexual stage is known for
E. parva
, the type species of the genus
Emmonsia
. To evaluate relationships among members of the putative
Ajellomyces
clade, large-subunit ribosomal and internal transcribed spacer region DNA sequences were determined from PCR-amplified DNA fragments. Sequences were analyzed phylogenetically to evaluate the genetic variation within the genus
Emmonsia
and evolutionary relationships to other taxa.
E. crescens
and
E. parva
are distinct species.
E. crescens
isolates are placed into two groups that correlate with their continents of origin. Considerable variation occurred among isolates previously classified as
E. parva
. Most isolates are placed into two closely related groups, but the remaining isolates, including some from human sources, are phylogenetically distinct and represent undescribed species. Strains of
B. dermatitidis
are a sister species of
E. parva
.
Paracoccidioides brasiliensis
and
Histoplasma capsulatum
are ancestral to most
Emmonsia
isolates, and
P. brasiliensis
, which has no known teleomorph, falls within the
Ajellomyces
clade.
Genetic and Serological Evidence for Multiple Instances of Unrecognized Transmission of Hepatitis C Virus in Hemodialysis Units
Authors: Mizuno Mari , Higuchi Terumi , Kanmatsuse Katsuo , Esumi Mariko 
Published: December 2019

ABSTRACT
We investigated the unrecognized patient-to-patient transmission of hepatitis C virus (HCV) in hemodialysis units by performing phylogenetic and serological analyses of hypervariable region 1 (HVR1) of HCV. Of the 62 patients in one center, 11 were positive for HCV RNA. A total of 24 HVR1 sequences, including the minor population of sequences of HCV isolates, from each patient were closely related and classified into five clusters by phylogenetic analysis. Of the 11 patients, 5 were infected with multiple clusters of HCV. Two patients were infected with HCV during an 18-month interval between examinations, and these HVR1 sequences fell into one of the five clusters. In another hemodialysis center, 5 of the 20 patients were HCV RNA positive, and two HVR1 sequences were found to be closely related and phylogenetically derived from the same cluster. The antibody responses of these patients to the HVR1 peptides representative of the genetic clusters revealed exactly the same clustering as that shown by phylogenetic analysis. These findings suggest that phylogenetic and serological analyses of HVR1 sensitively detect unrecognized and multiple transmission of HCV occurring within the same room in hemodialysis centers. Fingerprinting analyses using hypervariable regions of infectious agents are useful in identifying the precise route of transmission of infection.
Comparison of Restriction Enzyme Analysis, Arbitrarily Primed PCR, and Protein Profile Analysis Typing for Epidemiologic Investigation of an Ongoing
Clostridium difficile
Outbreak
Authors: Rafferty Mary Ellen , Baltch Aldona L. , Smith Raymond P. , Bopp Lawrence H. , Rheal Carol , Tenover Fred C. , Killgore George E. , Lyerly David M. , Wilkins Tracy D. , Schoonmaker Dianna J. , Hannett George E. , Shayegani Mehdi 
Published: December 2019

ABSTRACT

During an outbreak of diarrhea in a general hospital in 1992, 166
Clostridium difficile
isolates from 102 patients were typed by restriction enzyme analysis (REA), arbitrarily primed PCR (AP-PCR), and protein profile analysis (PP) techniques. A total of 18 types and 5 subtypes were identified by REA, 32 types were identified by AP-PCR, and 9 types were identified by PP. Analysis of the data indicated the presence of a predominant strain among 76, 75, and 84% of the isolates by REA, AP-PCR, and PP, respectively. Subsequently, 45
C. difficile
isolates which had been collected in 1990 from 33 patients in the same hospital following a significant increase in the number of cases of diarrhea caused by
C. difficile
were studied by REA, AP-PCR, and PP typing techniques. Thirteen types and one subtype were identified by REA, 12 types were identified by AP-PCR, and 5 types were identified by PP. As with the isolates from 1992, a dominant strain was identified. This strain was represented by 53, 64, and 70% of the total number of isolates when the strains were typed by REA, AP-PCR, and PP, respectively. Every isolate (210 of 211) from both 1990 and 1992 that was available for typing was typeable by all three methods. Furthermore, the same dominant strain was identified in both 1990 and 1992 by each method. This study demonstrates that each of the three typing methods can be useful in epidemiologic investigations of
C. difficile
outbreaks and that one strain can be dominant in an institution over a number of years.
Assessment of Hepatitis C Virus Sequence Complexity by Electrophoretic Mobilities of Both Single-and Double-Stranded DNAs
Authors: Wang Yu-Ming , Ray Stuart C. , Laeyendecker Oliver , Ticehurst John R. , Thomas David L. 
Published: December 2019

ABSTRACT

To assess genetic variation in hepatitis C virus (HCV) sequences accurately, we optimized a method for identifying distinct viral clones without determining the nucleotide sequence of each clone. Twelve serum samples were obtained from seven individuals soon after they acquired HCV during a prospective study, and a 452-bp fragment from the E2 region was amplified by reverse transcriptase PCR and cloned. Thirty-three cloned cDNAs representing each specimen were assessed by a method that combined heteroduplex analysis (HDA) and a single-stranded conformational polymorphism (SSCP) method to determine the number of clonotypes (electrophoretically indistinguishable cloned cDNAs) as a measure of genetic complexity (this combined method is referred to herein as the HDA+SSCP method). We calculated Shannon entropy, incorporating the number and distribution of clonotypes into a single quantifier of complexity. These measures were evaluated for their correlation with nucleotide sequence diversity. Blinded analysis revealed that the sensitivity (ability to detect variants) and specificity (avoidance of false detection) of the HDA+SSCP method were very high. The genetic distance (mean ± standard deviation) between indistinguishable cloned cDNAs (intraclonotype diversity) was 0.6% ± 0.9%, and 98.7% of cDNAs differed by <2%, while the mean distance between cloned cDNAs with different patterns was 4.0% ± 3.2%. The sensitivity of the HDA+SSCP method compared favorably with either HDA or the SSCP method alone, which resulted in intraclonotype diversities of 1.6% ± 1.8% and 3.5% ± 3.4%, respectively. The number of clonotypes correlated strongly with genetic diversity (
R
2
, 0.93), but this correlation fell off sharply when fewer clones were assessed. This HDA+SSCP method accurately reflected nucleotide sequence diversity among a large number of viral cDNA clones, which should enhance analyses to determine the effects of viral diversity on HCV-associated disease. If sequence diversity becomes recognized as an important parameter for staging or monitoring of HCV infection, this method should be practical enough for use in laboratories that perform nucleic acid testing.