bluuebunny/crossref_metadata_2025_split huggingface dataset:
Detection and Quantitation of Human Papillomavirus by Using the Fluorescent 5′ Exonuclease Assay
Authors: Josefsson Agnetha , Livak Ken , Gyllensten Ulf 
Published: December 2019

ABSTRACT

A method for the detection and quantitation of oncogenic human papillomavirus (HPV) was developed by using the fluorescent 5′ exonuclease assay. The method is based on the amplification of a 180-bp fragment from the 3′ part of the E1 open reading frame in a single PCR with type-specific probes for HPV types 16, 18, 31, 33, and 35. The probes can be used separately or in combinations of up to three probes per assay. Quantitation over a range of 10
1
to 10
6
initial HPV copies was possible by using real-time detection of the accumulation of fluorescence with cycle number. Reconstitution experiments, performed to mimic mixed infections, showed that individual HPV types can be detected down to a ratio of about 1% in a mixture. The performance of the assay depends on DNA quality, the presence of PCR inhibitors, and the number of different probes used simultaneously. This homogeneous assay provides a fast and sensitive way of screening for oncogenic HPV types in biopsy specimens as well as cervical smear samples. The closed-tube nature of the assay and the inclusion of uracil
N
′-glycosylase reduces cross contamination of PCR products to a minimum. A similar assay for β-actin was used in parallel for quantitation of genomic DNA. After normalizing the samples for genomic DNA content, the mean number of HPV copies per cell could be calculated.
Improved Detection of Rhinoviruses in Clinical Samples by Using a Newly Developed Nested Reverse Transcription-PCR Assay
Authors: Andeweg Arno C. , Bestebroer Theo M. , Huybreghs Martijn , Kimman Tjeerd G. , de Jong Jan C. 
Published: December 2019

ABSTRACT
This paper describes the development and evaluation of a new nested reverse transcription (RT)-PCR for the detection of rhinovirus in clinical samples. The nucleotide sequences of the 5′ noncoding regions of 39 rhinoviruses were determined in order to map the most conserved subregions. We designed a set of rhinovirus-specific primers and probes directed to these subregions and developed a new nested RT-PCR. The new assay includes an optimal RNA extraction method and amplicon identification with probe hybridization to discriminate between rhinoviruses and the closely related enteroviruses. It proved to be highly sensitive and specific. When tested on a dilution series of cultured viruses, the new PCR protocol scored positive at 10- to 100-fold-higher dilutions than a previously used nested RT-PCR. When tested on a collection of clinical samples obtained from 1,070 acute respiratory disease patients who had consulted their general practitioners, the new assay demonstrated a rhinovirus in 24% of the specimens, including all culture-positive samples, whereas the previously used PCR assay or virus culture detected a rhinovirus in only 3.5 to 6% of the samples. This new assay should help determine the disease burden associated with rhinovirus infections.
Trends in Antifungal Use and Epidemiology of Nosocomial Yeast Infections in a University Hospital
Authors: Berrouane Yasmina F. , Herwaldt Loreen A. , Pfaller Michael A. 
Published: December 2019

ABSTRACT

This report describes both the trends in antifungal use and the epidemiology of nosocomial yeast infections at the University of Iowa Hospitals and Clinics between fiscal year (FY) 1987–1988 and FY 1993–1994. Data were gathered retrospectively from patients’ medical records and from computerized databases maintained by the Pharmacy, the Program of Hospital Epidemiology, and the Medical Records Department. After fluconazole was introduced, use of ketoconazole decreased dramatically but adjusted use of amphotericin B decreased only moderately. However, the proportion of patients receiving antifungal therapy who were treated with amphotericin B declined markedly. In FY 1993–1994, 26 patients of the gastrointestinal surgery service received fluconazole. Among these patients, fluconazole use was prophylactic in 16 (61%), empiric in 3 (12%), and directed to a documented fungal infection in 7 (27%). Rates of nosocomial yeast infection in the adult bone marrow transplant unit increased from 6.77/1,000 patient days in FY 1987–1988 to 10.18 in FY 1989–1990 and then decreased to 0 in FY 1992–1993. Rates of yeast infections increased threefold in the medical and surgical intensive care units, reaching rates in FY 1993–1994 of 6.95 and 5.25/1,000 patient days, respectively. The rate of bloodstream infections increased from 0.044/1,000 patient days to 0.098, and the incidence of catheter-related urinary tract infections increased from 0.23/1,000 patient days to 0.68. Although the proportion of infections caused by yeast species other than
Candida albicans
did not increase consistently,
C. glabrata
became an important nosocomial pathogen.
Comparison of Agar Dilution, Disk Diffusion, MicroScan, and Vitek Antimicrobial Susceptibility Testing Methods to Broth Microdilution for Detection of Fluoroquinolone-Resistant Isolates of the Family
Enterobacteriaceae
Authors: Steward Christine D. , Stocker Sheila A. , Swenson Jana M. , O’Hara Caroline M. , Edwards Jonathan R. , Gaynes Robert P. , McGowan John E. , Tenover Fred C. 
Published: December 2019

ABSTRACT

Fluoroquinolone resistance appears to be increasing in many species of bacteria, particularly in those causing nosocomial infections. However, the accuracy of some antimicrobial susceptibility testing methods for detecting fluoroquinolone resistance remains uncertain. Therefore, we compared the accuracy of the results of agar dilution, disk diffusion, MicroScan Walk Away Neg Combo 15 conventional panels, and Vitek GNS-F7 cards to the accuracy of the results of the broth microdilution reference method for detection of ciprofloxacin and ofloxacin resistance in 195 clinical isolates of the family
Enterobacteriaceae
collected from six U.S. hospitals for a national surveillance project (Project ICARE [Intensive Care Antimicrobial Resistance Epidemiology]). For ciprofloxacin, very major error rates were 0% (disk diffusion and MicroScan), 0.9% (agar dilution), and 2.7% (Vitek), while major error rates ranged from 0% (agar dilution) to 3.7% (MicroScan and Vitek). Minor error rates ranged from 12.3% (agar dilution) to 20.5% (MicroScan). For ofloxacin, no very major errors were observed, and major errors were noted only with MicroScan (3.7% major error rate). Minor error rates ranged from 8.2% (agar dilution) to 18.5% (Vitek). Minor errors for all methods were substantially reduced when results with MICs within ±1 dilution of the broth microdilution reference MIC were excluded from analysis. However, the high number of minor errors by all test systems remains a concern.
Multicenter Comparison of the Sensititre YeastOne Colorimetric Antifungal Panel with the National Committee for Clinical Laboratory Standards M27-A Reference Method for Testing Clinical Isolates of Common and Emerging
Candida
spp.,
Cryptococcus
spp., and Other Yeasts and Yeast-Like Organisms
Authors: Espinel-Ingroff A. , Pfaller M. , Messer S. A. , Knapp C. C. , Killian S. , Norris H. A. , Ghannoum M. A. 
Published: December 2019

ABSTRACT

National Committee for Clinical Laboratory Standards (NCCLS) standard guidelines are available for the antifungal susceptibility testing of common
Candida
spp. and
Cryptococcus neoformans
, but NCCLS methods may not be the most efficient and convenient procedures for use in the clinical laboratory. MICs of amphotericin B, fluconazole, flucytosine, itraconazole, and ketoconazole were determined by the commercially prepared Sensititre YeastOne Colorimetric Antifungal Panel and by the NCCLS M27-A broth microdilution method for 1,176 clinical isolates of yeasts and yeast-like organisms, including
Blastoschizomyces capitatus
,
Cryptococcus
spp., 14 common and emerging species of
Candida
,
Hansenula anomala
,
Rhodotorula
spp.,
Saccharomyces cerevisiae
,
Sporobolomyces salmonicolor
, and
Trichosporon beigelii
. Colorimetric MICs of amphotericin B corresponded to the first blue well (no growth), and MICs of the other agents corresponded to the first purple or blue well. Three comparisons of MIC pairs by the two methods were evaluated to obtain percentages of agreement: 24- and 48-h MICs and 24-h colorimetric versus 48-h reference MICs. The best performance of the YeastOne panel was with 24-h MICs (92 to 100%) with the azoles and flucytosine for all the species tested, with the exception of
C. albicans
(87 to 90%). For amphotericin B, the best agreement between the methods was with 48-h MIC pairs (92 to 99%) for most of the species tested. The exception was for isolates of
C. neoformans
(76%). These data suggest the potential value of the YeastOne panel for use in the clinical laboratory.
Mycoplasma hyopneumoniaePotentiation of Porcine Reproductive and Respiratory Syndrome Virus-Induced Pneumonia
Authors: Thacker Eileen L. , Halbur Patrick G. , Ross Richard F. , Thanawongnuwech Roongroje , Thacker Brad J. 
Published: December 2019

ABSTRACTAn experimental model that demonstrates a mycoplasma species acting to potentiate a viral pneumonia was developed.Mycoplasma hyopneumoniae, which produces a chronic, lymphohistiocytic bronchopneumonia in pigs, was found to potentiate the severity and the duration of a virus-induced pneumonia in pigs. Pigs were inoculated withM. hyopneumoniae21 days prior to, simultaneously with, or 10 days after inoculation with porcine reproductive and respiratory syndrome virus (PRRSV), which induces an acute interstitial pneumonia in pigs. PRRSV-induced clinical respiratory disease and macroscopic and microscopic pneumonic lesions were more severe and persistent inM. hyopneumoniae-infected pigs. At 28 or 38 days after PRRSV inoculation,M. hyopneumoniae-infected pigs still exhibited lesions typical of PRRSV-induced pneumonia, whereas the lungs of pigs which had received only PRRSV were essentially normal. On the basis of macroscopic lung lesions, it appears that PRRSV infection did not influence the severity ofM. hyopneumoniaeinfection, although microscopic lesions typical ofM. hyopneumoniaewere more severe in PRRSV-infected pigs. These results indicate thatM. hyopneumoniaeinfection potentiates PRRSV-induced disease and lesions. Most importantly,M. hyopneumoniae-infected pigs with minimal to nondetectable mycoplasmal pneumonia lesions manifested significantly increased PRRSV-induced pneumonia lesions compared to pigs infected with PRRSV only. This discovery is important with respect to the control of respiratory disease in pigs and has implications in elucidating the potential contribution of mycoplasmas in the pathogenesis of viral infections of other species, including humans.
Pneumolysin PCR-Based Diagnosis of Invasive Pneumococcal Infection in Children
Authors: Toikka Pia , Nikkari Simo , Ruuskanen Olli , Leinonen Maija , Mertsola Jussi 
Published: December 2019

ABSTRACT

Blood-based pneumolysin PCR was compared to blood culture and detection of pneumolysin immune complexes, as well as to detection of antibodies to pneumolysin and to C polysaccharide, in the diagnosis of pneumococcal infection in 75 febrile children. Invasive pneumococcal infection was suspected on clinical grounds in 67 of the febrile children, and viral infection was suspected on clinical grounds in 8 of the febrile children. In addition, 15 healthy persons were examined to test the specificity of the PCR assay. Plasma, serum, and leukocyte fractions were analyzed by PCR. The combination of all test results led to the diagnosis of pneumococcal infection in 25 patients. Pneumolysin PCR was positive in 44% of these children, an increase occurred in the pneumolysin antibodies in 39% and in the C polysaccharide antibodies in 30% of the patients; pneumolysin immune complexes were found in convalescent serum in 30%, pneumolysin immune complexes occurred in acute-phase serum samples in 16%, and a positive blood culture was found in 20% of the patients. None of the healthy controls had positive results by PCR. The results suggest that the diagnosis of
Streptococcus pneumoniae
infection from blood samples necessitates the use of several different assays. Pneumolysin PCR was the most sensitive assay, but its clinical value is reduced by the fact that three blood fractions are needed.
Validation of Binary Typing for
Staphylococcus aureus
Strains
Authors: van Leeuwen Willem , Verbrugh Henri , van der Velden Jos , van Leeuwen Nan , Heck Max , van Belkum Alex 
Published: December 2019

ABSTRACT

Most of the DNA-based methods for genetic typing of
Staphylococcus aureus
strains generate complex banding patterns. Therefore, we have developed a binary typing procedure involving strain-differentiating DNA probes which were generated on the basis of randomly amplified polymorphic DNA (RAPD) analysis. We present and validate the usefulness of 15 DNA probes, according to generally accepted performance criteria for molecular typing systems. RAPD analysis with multiple primers was performed on 376
S. aureus
strains of which 97% were methicillin resistant (MRSA). Among the 1,128 RAPD patterns generated, 66 were selected which identified 124 unique DNA fragments. From these amplicons, only 12% turned out to be useful for isolate-specific binary typing. The nature of the RAPD-generated DNA fragments was investigated by partial DNA sequence analysis. Several homologies with known
S. aureus
sequences and with genes from other species were discovered; however, 87% of the probe sequences are of previously unknown origin. The locations of most of the DNA probes on the chromosome of
S. aureus
NCTC 8325 were determined by hybridization. Seven fragments were randomly dispersed along the genome, five were clustered within the 2500- to 2600-kb position of the genome, and the remaining four did not recognize complementary sequences in
S. aureus
NCTC 8325. A total of 103
S. aureus
strains (69% MRSA) were used for the validation of the binary typing technique. The 15 DNA probes provided stable epidemiological markers, both in vitro (type consistency after serial passages on culture media) and in vivo (comparison of sequential isolates recovered from cases of persistent colonization). The discriminatory power of binary typing (
D
= 0.998) exceeded that of pulsed-field gel electrophoresis (
D
= 0.966) and RAPD analysis (
D
= 0.949). Reproducibility, measured by analyzing multiple strains belonging to a multitude of different epidemiological clusters, was comparable to that of other genotyping techniques used. Contribution of the DNA probes to the discriminatory power of the system was analyzed by comparison of dendrograms. This study demonstrates that binary typing is a robust tool for the genetic typing of
S. aureus
isolates.
PCR Detection of Adenovirus in a Bone Marrow Transplant Recipient: Hemorrhagic Cystitis as a Presenting Manifestation of Disseminated Disease
Authors: Echavarria Marcela S. , Ray Stuart C. , Ambinder Richard , Dumler J. Stephen , Charache Patricia 
Published: December 2019

ABSTRACT
Adenoviruses (AdV), causing fatal disseminated infections in bone marrow transplant (BMT) recipients, are associated not only with hemorrhagic cystitis (HC) but also with hepatitis, conjunctivitis, and viral interstitial pneumonia. The importance of this virus as a cause of disseminated disease, however, has remained underappreciated. AdV infection has been diagnosed primarily through the use of cell culture. The fact that cell culture is insensitive for detecting this virus has hindered recognition of the role that AdV may play in morbidity and mortality in BMT recipients. To emphasize these points, we describe a patient who presented with HC due to AdV serotype 11, genotype c, and died with disseminated infection. In addition to cell culture, this study used a newly developed PCR-based method, capable of detecting all AdV serotypes tested, including different genotypes of serotype 11. The PCR result was positive in all culture-positive samples, including samples of urine, conjunctiva, and bronchoalveolar lavage (BAL). Importantly, the PCR method provided evidence of urinary shedding of AdV in a pretransplant, culture-negative specimen and showed dissemination in a subset of culture-negative specimens, including BAL, blood, and bone marrow samples. The lack of widespread awareness of the fact that localized infections may presage dissemination, and the previous associated lack of rapid, sensitive diagnostic assays, has impaired recognition of AdV infections in patients undergoing BMT. Early detection may contribute to therapy modification and avoidance of unwarranted diagnostic procedures. It may also assist in epidemiologic control of this highly infectious pathogen and lead to a renewed interest in preventive and therapeutic approaches.
In Vitro Culture and Drug Sensitivity Assay of
Plasmodium falciparum
with Nonserum Substitute and Acute-Phase Sera
Authors: Ringwald Pascal , Meche Fleurette Solange , Bickii Jean , Basco Leonardo K. 
Published: December 2019

ABSTRACT

The short-term in vitro growth of
Plasmodium falciparum
parasites in the asexual erythrocytic stage and the in vitro activities of eight standard antimalarial drugs were assessed and compared by using RPMI 1640 medium supplemented with 10% nonimmune human serum, 10% autologous or homologous acute-phase serum, or 0.5% Albumax I (lipid-enriched bovine serum albumin). In general, parasite growth was maximal with autologous (or homologous) serum, followed by Albumax I and nonimmune serum. The 50% inhibitory concentrations (IC
50
s) varied widely, depending on the serum or serum substitute. The comparison of IC
50
s between assays with autologous and nonimmune sera showed that monodesethylamodiaquine, halofantrine, pyrimethamine, and cycloguanil had similar IC
50
s. Although the IC
50
s of chloroquine, monodesethylamodiaquine, and dihydroartemisinin were similar with Albumax I and autologous sera, the IC
50
s of all test compounds obtained with Albumax I differed considerably from the corresponding values obtained with nonimmune serum. Our results suggest that Albumax I and autologous and homologous sera from symptomatic, malaria-infected patients may be useful alternative sources of serum for in vitro culture of
P. falciparum
isolates in the field. However, autologous sera and Albumax I do not seem to be suitable for the standardization of isotopic in vitro assays for all antimalarial drugs.
Comparative Evaluation of First- and Second-Generation Digene Hybrid Capture Assays for Detection of Human Papillomaviruses Associated with High or Intermediate Risk for Cervical Cancer
Authors: Poljak Mario , Brenčič Anja , Seme Katja , Vince Adriana , Marin Irena J. 
Published: December 2019

ABSTRACT
In the present study we comparatively evaluated the first- and second-generation Digene Hybrid Capture assays for detection of human papillomaviruses (HPV) associated with high or intermediate risk for cervical cancer in cervical specimens. Concordant results were obtained with 468 of 483 (96.8%) specimens. All 15 specimens which gave repeatedly discordant results were positive by the second-generation test only, and 14 of them tested PCR positive. The enhanced sensitivity of the second-generation assay is mainly a result of the reformulation of hybridization reagents and, to a lesser extent, a result of the addition of new HPV probes.
Antimicrobial Resistance and Serotype Distribution of
Streptococcus pneumoniae
Strains Causing Childhood Infections in Bangladesh, 1993 to 1997
Authors: Saha Samir K. , Rikitomi N. , Ruhulamin M. , Masaki H. , Hanif M. , Islam Maksuda , Watanabe K. , Ahmed K. , Matsumoto K. , Sack R. B. , Nagatake T. 
Published: December 2019

ABSTRACT

Three hundred sixty-two
Streptococcus pneumoniae
strains were isolated from children under 5 years of age at Dhaka Shishu (Children) Hospital from 1993 to 1997. The strains were isolated from blood (
n
= 105), CSF (
n
= 164), ear swab (
n
= 61), eye swab (
n
= 20), and pus (
n
= 12). Of the 362 isolates, 42 (11.6%) showed intermediate resistance (MIC, <0.1 μg/ml) and only 4 (1.1%) showed complete resistance (MIC, >2.0 μg/ml) to penicillin. Penicillin resistance exhibited a strong relationship with serotype 14; 47.8% of the penicillin-resistant strains belonged to this type. A remarkably high (64.1%) resistance to co-trimoxazole was observed, along with a significant increase during the time period studied; there was no relationship to capsular type. By way of contrast, penicillin resistance did not show any significant change during the study period. Resistance to chloramphenicol (2.2%) and erythromycin (1.1%) was rare. The high resistance to co-trimoxazole and its increasing trend demand elucidation of the clinical impact of pneumonia treatment by this antimicrobial and reconsideration of the World Health Organization recommendation for co-trimoxazole administration to children with community-acquired pneumonia at the health care worker level in Bangladesh.
Prevalence of Enterotoxigenic
Bacteroides fragilis
in Children with Diarrhea in Japan
Authors: Kato Naoki , Liu Chengxu , Kato Haru , Watanabe Kunitomo , Nakamura Haruhi , Iwai Naoichi , Ueno Kazue 
Published: December 2019

ABSTRACT

In age-matched controlled studies performed in Japan, enterotoxigenic
Bacteroides fragilis
was isolated from 14.9% of 114 children aged 1 to 14 years with antibiotic-unassociated diarrhea (AUD) and 6.5% of 108 children aged 1 to 6 years with antibiotic-associated diarrhea (AAD). The difference in comparison with control children, was significant for AUD children but not AAD children.
Justification for Use of a Single Trichrome Stain as the Sole Means for Routine Detection of Intestinal Parasites in Concentrated Stool Specimens
Authors: Kellogg James A. , Elder Carol J. 
Published: December 2019

ABSTRACT

Of 12,321 stool samples analyzed over a 6-year interval, 870 (7.1%) were positive for a total of 1,019 parasites, of which 1,011 (99.2%) were found in trichrome-stained smears of unconcentrated specimens while only 479 (47.0%) were detected in iodine-stained smears of concentrated samples. Stool specimens were next analyzed by trichrome staining of both unconcentrated and concentrated specimens preserved in either mercury-polyvinyl alcohol (PVA) or cupric PVA. Of 2,198 specimens, 171 (7.8%) were positive for a total of 208 parasites, 192 (92.3%) and 204 (98.1%) of which were found in the unconcentrated and concentrated specimens, respectively (
P
< 0.05). In our patient population, examination of a single trichrome-stained smear of a concentrated stool specimen is a cost-effective alternative to routinely analyzing both concentrated and unconcentrated specimens for parasites.
Evaluation of the Fungitest Kit by Using Strains from Human Immunodeficiency Virus-Infected Patients: Study of Azole Drug Susceptibility
Authors: Witthuhn F. , Toubas D. , Béguinot I. , Aubert D. , Rouger C. , Remy G. , Pinon J. M. 
Published: December 2019

ABSTRACT

One hundred eighteen
Candida
clinical isolates from human immunodeficiency virus-infected patients were tested for their susceptibilities to fluconazole and itraconazole by Fungitest and the National Committee for Clinical Laboratory Standards MIC method. Fungitest results depended on both yeast species and antifungal agents. This test is able to detect sensitive strains (97% agreement with results of the MIC method in tests with fluconazole and 84% agreement in tests with itraconazole) but has a poor capacity to detect resistant strains (26% agreement in tests with fluconazole and 5% agreement in tests with itraconazole).