bluuebunny/crossref_metadata_2025_split huggingface dataset:
Q Fever in the Greek Island of Crete: Detection, Isolation, and Molecular Identification of Eight Strains of
Coxiella burnetii
from Clinical Samples
Authors: Spyridaki Ioanna , Gikas Achilleas , Kofteridis Diamantis , Psaroulaki Anna , Tselentis Yannis 
Published: December 2019

ABSTRACT

Over a period of 6 years (1989 to 1995), serum samples from 3,300 patients suspected to be infected by
Coxiella burnetii
were assayed for the presence of antibodies against antigen phase II of the microorganism by the indirect immunofluorescence antibody technique (IFAT). One hundred fifty-two cases were recorded, and blood samples from 17 patients were cultured for the isolation of the pathogen. By a centrifugation shell vial technique, eight strains were isolated from patients suffering from acute Q fever. The microorganism was detected in the cultures by IFAT, by Gimenez staining, and by the cytopathogenic effect on Vero and human embryonic lung (HEL) cells. PCR followed by restriction fragment length polymorphism analysis was used to confirm the diagnosis and identify the
Coxiella burnetii
strains within the cell cultures as well as to compare them with reference strains. In order to avoid time-consuming cultures, to achieve direct detection of
Coxiella burnetii
in clinical samples (blood, buffy coat, etc.), and to increase the specificity and sensitivity of the detection, nested PCR was performed. The first step of DNA extraction was performed with the QIAamp blood kit 250. For the second step of the PCR assays, the conditions of temperature and times of recycling were properly modified, and the microorganism was detected within 4 h. Our study demonstrates that Q fever is an endemic disease in Crete and that the diagnosis of
Coxiella burnetii
infection can be rapidly achieved by the detection of the microorganism in buffy coat samples by nested PCR. Although the presenting symptoms of the disease in this study differed from those in other studies, the Cretan strains do not differ genotypically from the reference strains (Nine Mile and Q212).
Diagnostic Use of PCR for Detection of
Pneumocystis carinii
in Oral Wash Samples
Authors: Helweg-Larsen Jannik , Skov Jensen Jørgen , Benfield Thomas , Gerner Svendsen Ulrik , Lundgren Jens D. , Lundgren Bettina 
Published: December 2019

ABSTRACT

There is a need to develop noninvasive methods for the diagnosis of
Pneumocystis carinii
pneumonia in patients unable to undergo bronchoscopy or induction sputum. Oral wash specimens are easily obtained, and
P. ca- rinii
nucleic acid can be amplified and demonstrated by PCR. In routine clinical use, easy sample processing and single-round PCR are needed to ensure rapid analysis and to reduce the risk of contamination. We developed a single-round Touchdown PCR (TD-PCR) protocol with the ability to detect PCR inhibition in the specimen. The TD-PCR was evaluated in a routine diagnostic laboratory and was compared to a previously described PCR protocol (mitochondrial RNA) run in a research laboratory. Both PCR methods amplified a sequence of the mitochondrial rRNA gene of
P. carinii
. Paired bronchoalveolar lavage (BAL) and oral wash specimens from 76 consecutive human immunodeficiency virus type 1-infected persons undergoing a diagnostic bronchoscopy were included. The TD-PCR procedure was quicker than the mitochondrial PCR procedure (<24 versus 48 h) and, compared to microscopy, had sensitivity, specificity, and positive and negative predictive values of 89, 94, 93, and 91%, respectively, for oral wash specimens and 100, 91, 90, and 100%, respectively, for BAL specimens. Our results suggest that oral wash specimens are a potential noninvasive method to obtain a diagnostic specimen during
P. carinii
pneumonia infection and that it can be applied in a routine diagnostic laboratory.
Assessment of Spontaneous Fluctuations of Viral Load in Untreated Patients with Chronic Hepatitis C by Two Standardized Quantitation Methods: Branched DNA and Amplicor Monitor
Authors: Halfon Philippe , Bourlière Marc , Halimi Gilles , Khiri Hacène , Bertezene Patrice , Portal Isabelle , Botta-Fridlund Danielle , Gauthier AndréPierre , Jullien Monique , Feryn Jean Marc , Gerolami Victoria , Cartouzou Guy 
Published: December 2019

ABSTRACT
Quantitation of hepatitis C virus (HCV) RNA in serum has been used to predict and monitor the efficacy of interferon therapy in chronic HCV infection. We prospectively studied the fluctuation of viremia by a longitudinal follow-up of HCV RNA levels for 2 months in six untreated patients. Spontaneous fluctuations of HCV RNA ranged from 2.8- to 5.7-fold with branched DNA assay and from 2.9- to 5.6-fold with Monitor. These large spontaneous fluctuations (up to 0.75 log), observed daily, weekly, and monthly, raise doubt about the clinical value of a single assessment of pretherapeutic viremia.
Molecular Diagnosis of Human Rhinovirus Infections: Comparison with Virus Isolation
Authors: Hyypiä Timo , Puhakka Tuomo , Ruuskanen Olli , Mäkelä Mika , Arola Anita , Arstila Pertti 
Published: December 2019

ABSTRACT
To compare the sensitivity and specificity of RT-PCR with that of virus isolation in the detection of human rhinoviruses, we tested nasopharyngeal aspirates from 200 patients on the 1st and 7th days after the onset of the common cold. An assay utilizing a short amplicon in the conserved 5′ noncoding region was found highly sensitive. Of 192 positive samples altogether, 65 were found positive by RT-PCR only, 6 were positive by isolation exclusively, and 121 gave positive results in both tests.
Effects of Various Handling and Storage Conditions on Stability of
Treponema pallidum
DNA in Cerebrospinal Fluid
Authors: Villanueva A. V. , Podzorski R. P. , Reyes M. P. 
Published: December 2019

ABSTRACT

Treponema pallidum
DNA from even small numbers of organisms was detectable in cerebrospinal fluid (CSF) stored at room temperature or at 4°C for several hours and in CSF subjected to three freeze-thaw cycles. These results suggest that negative PCR results for
T. pallidum
from patients diagnosed with
T. pallidum
invasion of the central nervous system are probably not due to the loss of target DNA prior to testing.
Chromosomal Rearrangements in
Salmonella enterica
Serotype Typhi Affecting Molecular Typing in Outbreak Investigations
Authors: Echeita M. A. , Usera M. A. 
Published: December 2019

ABSTRACT

Salmonella enterica
serotype Typhi strains belonging to eight different outbreaks of typhoid fever that occurred in Spain between 1989 and 1994 were analyzed by ribotyping and pulsed-field gel electrophoresis. For three outbreaks, two different patterns were detected for each outbreak. The partial digestion analysis by the intron-encoded endonuclease I-
Ceu
I of the two different strains from each outbreak provided an excellent tool for examining the organization of the genomes of epidemiologically related strains.
S. enterica
serotype Typhi seems to be more susceptible than other serotypes to genetic rearrangements produced by homologous recombinations between
rrn
operons; these rearrangements do not substantially alter the stability or survival of the bacterium. We conclude that genetic rearrangements can occur during the emergence of an outbreak.
Therapeutic Effect of Doxycycline in Experimental Subclinical Canine Monocytic Ehrlichiosis: Evaluation of a 6-Week Course
Authors: Harrus Shimon , Waner Trevor , Aizenberg Itzhak , Bark Hylton 
Published: December 2019

ABSTRACTThe efficacy of doxycycline treatment (10 mg/kg of body weight every 24 h for 42 days) in eliminatingEhrlichia canisfrom four subclinically infected dogs was evaluated. One dog remained PCR positive, suggesting that 6 weeks of doxycycline treatment may not be sufficient to clearE. canisparasites from all subclinically infected dogs. Serology (indirect immunofluorescent antibody assay) was shown to be unreliable in assessing recovery from the carrier state, as anti-E. canisantibodies persisted after elimination of the parasite. Our findings suggest that an increase in the platelet count may be an important indicator for dogs that recover from subclinical ehrlichiosis.
Facklamia ignava
sp. nov., Isolated from Human Clinical Specimens
Authors: Collins Matthew D. , Lawson Paul A. , Monasterio Rafael , Falsen Enevold , Sjöden Berit , Facklam Richard R. 
Published: December 2019

ABSTRACT

Two strains of a hitherto-undescribed gram-positive, catalase-negative coccus isolated from human sources were characterized by phenotypic and molecular taxonomic methods. Comparative 16S rRNA gene sequencing studies demonstrated that the unknown strains are genealogically identical and constitute a new line close to, but distinct from,
Facklamia hominis
. The unknown bacterium was readily distinguished from
F. hominis
by biochemical tests and electrophoretic analysis of whole-cell proteins. On the basis of phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium be classified as
Facklamia ignava
sp. nov. The type strain of
Facklamia ignava
is CCUG 37419.
Diagnosis of Meningococcal Meningitis by Broad-Range Bacterial PCR with Cerebrospinal Fluid
Authors: Kotilainen Pirkko , Jalava Jari , Meurman Olli , Lehtonen Olli-Pekka , Rintala Esa , Seppälä Olli-Pekka , Eerola Erkki , Nikkari Simo 
Published: December 2019

ABSTRACT

We used broad-range bacterial PCR combined with DNA sequencing to examine prospectively cerebrospinal fluid (CSF) samples from patients with suspected meningitis. Fifty-six CSF samples from 46 patients were studied during the year 1995. Genes coding for bacterial 16S and/or 23S rRNA genes could be amplified from the CSF samples from five patients with a clinical picture consistent with acute bacterial meningitis. For these patients, the sequenced PCR product shared 98.3 to 100% homology with the
Neisseria meningitidis
sequence. For one patient, the diagnosis was initially made by PCR alone. Of the remaining 51 CSF samples, for 50 (98.0%) samples the negative PCR findings were in accordance with the negative findings by bacterial culture and Gram staining, as well as with the eventual clinical diagnosis for the patient. However, the PCR test failed to detect the bacterial rRNA gene in one CSF sample, the culture of which yielded
Listeria monocytogenes
. These results invite new research efforts to be focused on the application of PCR with broad-range bacterial primers to improve the etiologic diagnosis of bacterial meningitis. In a clinical setting, Gram staining and bacterial culture still remain the cornerstones of diagnosis.
Detection and Semiquantitative Analysis of Human Herpesvirus 8 DNA in Specimens from Patients with Kaposi’s Sarcoma
Authors: Mendez J. C. , Procop G. W. , Espy M. J. , Paya C. V. , Smith T. F. 
Published: December 2019

ABSTRACT
Kaposi’s sarcoma (KS) is the most common neoplasm in patients with AIDS. Epidemiologic evidence and the recent identification of herpesvirus-like DNA sequences in patients with KS have suggested a role for viral agents in the etiopathogenesis of this disease. It is unclear if these sequences are present in all types of KS and if the copy number of these sequences has a correlation with disease severity (staging). In order to clarify these issues, we retrospectively analyzed, by PCR and Southern blotting, formalin-fixed, paraffin-embedded biopsy specimens from 12 patients previously diagnosed with KS by histopathologic examination of these specimens between the years of 1977 and 1996. We also analyzed tissue samples from these patients taken from dermal sites without KS lesions and control tissues from healthy subjects. Of the 12 patients, 6 had classic KS, 5 had AIDS-associated KS, and 1 had the endemic type of KS. We tested the specimens for other herpesviruses, including cytomegalovirus, human herpesvirus 6 (HHV-6), HHV-7, HHV-8, and Epstein-Barr virus. Of 20 biopsy specimens from patients previously diagnosed with KS, 19 were positive for HHV-8 sequences (95%), while PCR for the DNAs of other herpesvirus agents was negative. Uninvolved tissue from these patients and control tissue from healthy subjects gave negative results for all viruses. Semiquantitative analysis of Southern blots showed higher levels of HHV-8 DNA in those patients with multicentric and visceral involvement than in those patients with localized involvement. In addition, in patients with localized skin disease, the nodular stage had higher levels of HHV-8 DNA than the patch or plaque stages. Our data confirmed that HHV-8 is involved in the etiopathogenesis of all types of KS and that there is a correlation between HHV-8 DNA load and the severity and staging of this disease.
Canine Intestinal Spirochetes Consist of
Serpulina pilosicoli
and a Newly Identified Group Provisionally Designated “
Serpulina canis
” sp. nov
Authors: Duhamel G. E. , Trott D. J. , Muniappa N. , Mathiesen M. R. , Tarasiuk K. , Lee J. I. , Hampson D. J. 
Published: December 2019

ABSTRACT

The spirochetes inhabiting the large intestines of humans and animals consist of a diverse group of related organisms. Intestinal spirochetosis caused by
Serpulina pilosicoli
is a newly recognized enteric disease of human beings and animals with potential public health significance. The purpose of this study was to determine the species identity of canine intestinal spirochetes by comparing 30 isolates obtained from dogs in Australia (
n
= 25) and the United States (
n
= 5) with reference strains representing
Serpulina
species and
Brachyspira aalborgi
, by phenotypic and genetically based typing methods. All of the canine isolates were indole negative and produced a weak β-hemolysis when cultured anaerobically on agar medium containing blood. Four isolates were identified as
S. pilosicoli
by 16S rRNA-specific PCR assays, rRNA gene restriction fragment length polymorphism or ribotyping, and multilocus enzyme electrophoresis. The remaining 26 isolates formed a cluster related to porcine
Serpulina innocens
as determined by multilocus enzyme electrophoresis but had a unique ribotype pattern. The data suggested the existence of a novel
Serpulina
species, provisionally designated “
Serpulina canis
,” colonizing the intestines of dogs.
Localization of Persistent
Enterocytozoon bieneusi
Infection in Normal Rhesus Macaques (
Macaca mulatta
) to the Hepatobiliary Tree
Authors: Mansfield Keith G. , Carville Angela , Hebert Daniel , Chalifoux Laura , Shvetz Daniel , Lin K. C. , Tzipori S. , Lackner Andrew A. 
Published: December 2019

ABSTRACT

Enterocytozoon bieneusi
is the most common microsporidian parasite recognized in human patients with AIDS. Recently, we identified a virtually identical organism causing a spontaneous infection associated with hepatobiliary and intestinal disease in simian immunodeficiency virus (SIV)-infected macaques. To examine the natural history of the infection, we examined captive rhesus macaques for
E. bieneusi
by PCR, in situ hybridization, and cytochemical techniques. PCR performed on fecal DNA detected enterocytozoon infection in 22 (16.7%) of 131 normal rhesus macaques (
Macaca mulatta
), compared to 18 (33.8%) of 53 rhesus macaques experimentally inoculated with SIV. In normal rhesus macaques, persistence of infection was demonstrated for up to 262 days and was usually not associated with clinical signs. In six of seven normal rhesus animals,
E. bieneusi
was detected by PCR in bile obtained through percutaneous cholecystocentesis but not by in situ hybridization performed on endoscopic biopsies of duodenum and proximal jejunum.
Efficient Isolation of Human Immunodeficiency Virus Type 1 RNA from Cervical Swabs
Authors: Hajjar Adeline M. , Lewis Paul F. , Endeshaw Yohannes , Ndinya-Achola Jackoniah , Kreiss Joan K. , Overbaugh Julie 
Published: December 2019

ABSTRACT

An efficient method for the isolation of human immunodeficiency virus type 1 (HIV-1) nucleic acids from dry cervical swabs was developed. HIV-1
gag
and
env
were detected in 96% (25 of 26) and 81% (21 of 26), respectively, of the samples tested by PCR from HIV-1-seropositive women in a Kenyan cohort study. Eighty-eight percent of the swabs (22 of 25) were positive for
gag
RNA, and 85% (17 of 20) were positive for
env
RNA. Fewer than 1,000 copies of HIV-1
gag
RNA were detected in four swabs in which a competitive quantitative PCR assay was used. The method described here may be useful for both qualitative and quantitative analyses of HIV RNA in mucosal secretions as well as amplification and cloning of full-length viral genes for functional studies.
In Vitro Testing of Susceptibilities of Filamentous Ascomycetes to Voriconazole, Itraconazole, and Amphotericin B, with Consideration of Phylogenetic Implications
Authors: McGinnis Michael R. , Pasarell Lester 
Published: December 2019

ABSTRACT
The in vitro susceptibilities of three hundred eighty-one isolates representing two classes, five orders, nine families, 30 genera, and 51 species of ascomycetous fungi to voriconazole, itraconazole, and amphotericin B were tested by using a modification of the National Committee for Clinical Laboratory Standards M27-A reference method. For those fungi of known phylogenetic relatedness, drug MICs were consistently low for isolates among all clades, except for members of the family Microascaceae. The highest MICs of all drugs tested were consistently for the Microascaceae, supporting the observation of fungal phylogeny and corresponding susceptibility to antifungal drugs. Itraconazole and voriconazole have a broad range of activity against phylogenetically similar agents of hyalohyphomycosis, phaeohyphomycosis, chromoblastomycosis, and mycetoma.
Human Infection with
Hymenolepis diminuta
: Case Report from Spain
Authors: Tena Daniel , Simón Montserrat Pérez , Gimeno Carmen , Pomata María Teresa Pérez , Illescas Soledad , Amondarain Izaskun , González Alejandro , Domínguez Jesús , Bisquert Julia 
Published: December 2019

ABSTRACT

We report a case of
Hymenolepis diminuta
infection in a human. The patient was a 5-year-old girl referred to us through the onset of a cyanotic attack. Treatment with a single dose (10 mg/kg of body weight) of praziquantel was ineffective, but the parasite was eradicated after three treatment cycles with the same drug at dosages of 25 mg/kg/day for 5 days.
Comparison of Broth Microdilution Method Using
Haemophilus
Test Medium and Agar Dilution Method for Susceptibility Testing of
Eikenella corrodens
Authors: Alcalá Luis , García-Garrote Fernando , Cercenado Emilia , Peláez Teresa , Ramos Gema , Bouza Emilio 
Published: December 2019

ABSTRACT

Susceptibility testing of
Eikenella corrodens
is usually performed by a Mueller-Hinton sheep blood agar dilution (AD) method. However, this method is impractical for testing only a few strains. We compared AD with the broth microdilution method using
Haemophilus
test medium (HTM) in order to determine the susceptibility of 36 clinical isolates of
E. corrodens
to eight antimicrobial agents. MICs obtained by the HTM method yielded 95.5 and 84% agreement (within 2 and 1 log
2
dilutions, respectively) with those obtained by AD. The HTM method with incubation in CO
2
for 48 h was highly reproducible and constitutes an easy alternative for antimicrobial susceptibility testing of
E. corrodens
.
Differentiation between
Mycobacterium tuberculosis
and
Mycobacterium avium
by Amplification of the 16S-23S Ribosomal DNA Spacer
Authors: Sansila Arunnee , Hongmanee Poonpilas , Chuchottaworn Charoen , Rienthong Somsak , Rienthong Dhanida , Palittapongarnpim Prasit 
Published: December 2019

ABSTRACT

Differentiation between
Mycobacterium tuberculosis
and
M. avium
is helpful for the treatment of disseminated mycobacterial infection in AIDS patients. This can traditionally be done by time-consuming biochemical tests or with Accuprobe. Previously, PCR restriction enzyme analysis (PCR-REA) of the 16S-23S rRNA gene spacer was shown to be able to identify a limited number of strains of
Mycobacterium
. In this study the method was improved by using more specific primers and was tested with 50 clinical isolates of
M. tuberculosis
and 65 clinical isolates of
M. avium
complex. Probes specific to the spacers of
M. tuberculosis
and
M. avium
were also tested. Both
M. tuberculosis
and
M. avium
could be reliably identified either by PCR-REA or by PCR-hybridization, with the results completely agreeing with those obtained by biochemical tests and with the Accuprobe, respectively. The method may therefore be useful as an alternative in-house method for identification of the bacteria.
Detection of Cytomegalovirus in Plasma and Cerebrospinal Fluid Specimens from Human Immunodeficiency Virus-Infected Patients by the AMPLICOR CMV Test
Authors: Long Christopher M. , Drew Lawrence , Miner Richard , Jekic-McMullen Dragana , Impraim Chaka , Kao Shaw-Yi 
Published: December 2019

ABSTRACT

We have developed the AMPLICOR CMV Test, which is rapid and sensitive for the detection of cytomegalovirus (CMV) in plasma and cerebrospinal fluid (CSF) specimens. The test incorporated an internal control in the reaction mixture to monitor the amplification efficiency and the presence of inhibitors. The AMPLICOR CMV Test was very specific in detecting 12 clinical CMV isolates and four laboratory CMV strains tested. Cross-reactivity with 26 non-CMV pathogens was not observed. The AMPLICOR CMV Test requires only 50 μl of specimen (plasma or CSF) for processing. The performance of the AMPLICOR CMV Test was compared to those of the CMV antigenemia assay and the conventional tube culture method. Among 112 plasma specimens from 43 human immunodeficiency virus-infected patients, CMV was detected in 20 (18%) of the specimens by the AMPLICOR CMV Test, 21 (19%) of the specimens by the CMV antigenemia assay, and 10 (9%) of the specimens by culture. In CSF specimens from AIDS patients, CMV was detected in 10 of 58 (17%) specimens tested by the AMPLICOR CMV Test, 5 of 28 (18%) specimens tested by the antigen assay, and none of the 25 specimens tested by culture. While the performance of the AMPLICOR CMV Test in this study was comparable to that of the CMV antigen assay, processing of specimens by the AMPLICOR CMV Test was much simpler than that by the antigen assay; in addition, the antigen assay requires greater than 10
5
leukocytes from blood or 1 ml of CSF to perform the assay. Our study suggested that the AMPLICOR CMV Test could provide a rapid and sensitive assay for the detection of CMV in plasma and CSF specimens.