bluuebunny/crossref_metadata_2025_split huggingface dataset:
Comparison of PCR, Nested PCR, and Random Amplified Polymorphic DNA PCR for Detection and Typing of
Ureaplasma urealyticum
in Specimens from Pregnant Women
Authors: Knox Christine L. , Timms Peter 
Published: December 2019

ABSTRACT

A PCR assay, using three primer pairs, was developed for the detection of
Ureaplasma urealyticum
, parvo biovar,
mba
types 1, 3, and 6, in cultured clinical specimens. The primer pairs were designed by using the polymorphic base positions within a 310- to 311-bp fragment of the 5′ end and upstream control region of the
mba
gene. The specificity of the assay was confirmed with reference serovars 1, 3, 6, and 14 and by the amplified-fragment sizes (81 bp for
mba
1, 262 bp for
mba
3, and 193 bp for
mba
6). A more sensitive nested PCR was also developed. This involved a first-step PCR, using the primers UMS-125 and UMA226, followed by the nested
mba
-type PCR described above. This nested PCR enabled the detection and typing of small numbers of
U. urealyticum
cells, including mixtures, directly in original clinical specimens. By using random amplified polymorphic DNA (RAPD) PCR with seven arbitrary primers, we were also able to differentiate the two biovars of
U. urealyticum
and to identify 13 RAPD-PCR subtypes. By applying these subtyping techniques to clinical samples collected from pregnant women, we established that (i)
U. urealyticum
is often a persistent colonizer of the lower genital tract from early midtrimester until the third trimester of pregnancy, (ii)
mba
type 6 was isolated significantly more often (
P
= 0.048) from women who delivered preterm than from women who delivered at term, (iii) no particular ureaplasma subtype(s) was associated with placental infections and/or adverse pregnancy outcomes, and (iv) the ureaplasma subtypes most frequently isolated from women were the same subtypes most often isolated from infected placentas.
Subcutaneous Hyalohyphomycosis Caused byColletotrichum gloeosporioides
Authors: Guarro Josep , Svidzinski Terezinha E. , Zaror Luís , Forjaz Maily H. , Gené Josepa , Fischman Olga 
Published: December 2019

ABSTRACTThe coelomyceteColletotrichum gloeosporioideswas isolated in pure culture from subcutaneous nodules of the left forearm and elbow of a farmer after traumatic injury. To our knowledge, we report the first case involving this fungus as an etiological agent of subcutaneous infection. The in vitro inhibitory activities of amphotericin B, itraconazole, ketoconazole, miconazole, flucytosine, and fluconazole were studied.
Detection of
Aspergillus
Galactomannan: Comparison of an Enzyme-Linked Immunoassay and a Europium-Linked Time-Resolved Fluoroimmunoassay
Authors: Paugam A. , Sarfati J. , Romieu R. , Viguier M. , Dupouy-Camet J. , Latgé J. P. 
Published: December 2019

ABSTRACT

With a view to improving the sensitivity of serological detection of
Aspergillus
galactomannan (GM), a europium-linked time-resolved fluoroimmunoassay was developed. This method was compared to an enzyme-linked immunosorbent assay using a peroxidase-conjugated detector antibody. No increase in the sensitivity of the detection of GM standards was seen with the europium-based fluoroimmunoassay.
Utility of Routine Testing of Bronchoalveolar Lavage Fluid for Cryptococcal Antigen
Authors: Kralovic Stephen M. , Rhodes Judith C. 
Published: December 2019

ABSTRACT
All cryptococcal antigen (CrAg) testing performed at our institution between 1989 and 1994 was reviewed for utility of routinely testing of bronchoalveolar lavage fluid (BAL) for this antigen. Forty-two of 1,506 BAL specimens were positive. Seventeen of these were felt to represent false positives (sensitivity, 71%; positive predictive value, 0.59). The data on CrAg in cerebrospinal fluid and serum and the fungal culture and histological results of BAL specimens did not support continued, routine testing of BALs for CrAg to diagnose cryptococcosis.
PCR-Based Rapid Detection of
Mycobacterium tuberculosis
in Blood from Immunocompetent Patients with Pulmonary Tuberculosis
Authors: Ahmed Niyaz , Mohanty Ashok Kumar , Mukhopadhyay Utpal , Batish Virender Kumar , Grover Sunita 
Published: December 2019

ABSTRACT

A PCR test based on insertion sequence IS1081 was developed to detect
Mycobacterium tuberculosis
complex organisms in the peripheral blood. The method was applied to blood samples from immunocompetent individuals with localized pulmonary tuberculosis. Seven of 16 (43.75%) blood samples were found to be positive for the circulating DNA copies of
M. tuberculosis
complex.
Extended-Spectrum β-Lactamase-Producing
Klebsiella pneumoniae
Strains Causing Nosocomial Outbreaks of Infection in the United Kingdom
Authors: Shannon Kevin , Stapleton Paul , Xiang Xiaoqin , Johnson Alan , Beattie Hamish , El Bakri Fatima , Cookson Barry , French Gary 
Published: December 2019

ABSTRACT

Representative isolates from 10 distinct extended-spectrum β-lactamase-producing strains of
Klebsiella pneumoniae
that caused hospital outbreaks in the United Kingdom from 1991 to 1994 were examined for relationships between their enzymes and plasmids. The β-lactamases were identified by a combination of isoelectric focusing and gene sequencing. SHV-2 β-lactamase was produced by isolates from four outbreaks, SHV-5 was involved in three, and SHV-4, TEM-15, and TEM-26 were involved in one outbreak each. All of the extended-spectrum β-lactamases were encoded by self-transmissible plasmids, with sizes ranging from about 70 to 160 kb. No similarities between the restriction digest patterns of the extended-spectrum β-lactamase-encoding plasmids were detected, except to some extent between those that produced TEM-15 and TEM-26. Thus, outbreaks of hospital infection with these organisms in the United Kingdom from 1991 to 1994 involved distinct organisms and resistance plasmids and appeared to be unrelated.
Fluorescent Amplified-Fragment Length Polymorphism Analysis of an Outbreak of Group A Streptococcal Invasive Disease
Authors: Desai Meeta , Tanna Asha , Wall Robert , Efstratiou Androulla , George Robert , Stanley John 
Published: December 2019

ABSTRACTFluorescent amplified-fragment length polymorphism (FAFLP) analysis was carried out for an outbreak of group A streptococcal (GAS) invasive disease. Streptococcal genomic DNAs were digested with endonucleasesEcoRI andMseI, site-specific adaptors were ligated, and PCR amplification was carried out with anEcoRI adaptor-specific primer labelled with fluorescent dye. Amplified fragments of up to 600 bp in size were separated on a polyacrylamide sequencing gel which contained internal size markers in each lane. These data were automatically scanned and analyzed, fragments were precisely sized (±1 bp), and electropherograms were generated for each genome with GeneScan 2.1 software. All isolates were compared in this way. Among 27 GAS isolates examined, we found 18 FAFLP profiles, compared with 12 macrorestriction profiles by pulsed-field gel electrophoresis. FAFLP readily distinguished genotypes for two clones of GAS serotype M77 which were responsible for outbreaks of invasive disease in a care-of-the-elderly system. It provided an automated analysis of the whole genome of bacterial isolates. It was reproducible, more discriminatory, and capable of higher throughput than other molecular typing methods. Given agreed conditions, FAFLP would be reproducible between laboratories for rapid characterization of outbreak strains.
Identification of
Legionella
Species by Random Amplified Polymorphic DNA Profiles
Authors: Lo Presti François , Riffard Serge , Vandenesch François , Etienne Jerome 
Published: December 2019

ABSTRACT

Random amplified polymorphic DNA (RAPD) was used for the identification of
Legionella
species. Primer SK2 (5′-CGGCGGCGGCGG-3′) and standardized RAPD conditions gave the technique a reproducibility of 93 to 100%, depending on the species tested. Species-specific patterns corresponding to the 42
Legionella
species were consequently defined by this method; the patterns were dependent on the recognition of a core of common bands for each species. This specificity was demonstrated by testing 65 type strains and 265 environmental and clinical isolates. No serogroup-specific profiles were obtained. A number of unidentified
Legionella
isolates potentially corresponding to new species were clustered in four groups. RAPD analysis appears to be a rapid and reproducible technique for identification of
Legionella
isolates to the species level without further restriction or hybridization.
Comparison of the MB/BacT System with a Revised Antibiotic Supplement Kit to the BACTEC 460 System for Detection of Mycobacteria in Clinical Specimens
Authors: Benjamin W. H. , Waites K. B. , Beverly A. , Gibbs L. , Waller M. , Nix S. , Moser S. A. , Willert M. 
Published: December 2019

ABSTRACT

The MB/BacT system (MB/BacT) with a revised antibiotic supplement kit was compared with the BACTEC 460 system (BACTEC 460) in a test of 488 specimens submitted for mycobacterial culture from 302 patients. Twenty-four
Mycobacterium tuberculosis
isolates were detected by the BACTEC 460 versus 23 isolates by the MB/BacT. Mean time until detection of
M. tuberculosis
isolates identified by both systems was 11.9 days for the BACTEC 460 versus 13.7 days for the MB/BacT (
P
= 0.046).
M. avium
complex was detected in 12 specimens by the MB/BacT versus 10 specimens by the BACTEC 460. Only 8 of 14 (57%)
M. avium
isolates were detected by both systems, with a mean time until detection of 10.1 days for the BACTEC 460 and 14.2 days for the MB/BacT (
P
= 0.009). The BACTEC 460 and the MB/BacT detected
M. gordonae
in four specimens, but only a single specimen was positive by both systems. One
M. fortuitum
isolate and one of five
M. kansasii
isolates were recovered only by the BACTEC 460. The bacterial overgrowth rate was 7.0% for the MB/BacT versus 4.1% for the BACTEC 460. We found the MB/BacT to be comparable to the BACTEC 460 for mycobacterial detection. Even though time until detection with the MB/BacT was slightly longer (1.8 days longer for
M. tuberculosis
and 4.1 days for
M. avium
[mean values]) and the bacterial overgrowth rate was somewhat higher, the decreased labor, the availability of a computerized data management system, and the noninvasive, nonradiometric aspects of the MB/BacT offset these relative disadvantages and make it an acceptable alternative for use in the diagnostic laboratory.
Prevalence of
Porphyromonas gingivalis
and Periodontal Health Status
Authors: Griffen Ann L. , Becker Mitzi R. , Lyons Sharon R. , Moeschberger Melvin L. , Leys Eugene J. 
Published: December 2019

ABSTRACT

Periodontitis is a common, progressive disease that eventually affects the majority of the population. The local destruction of periodontitis is believed to result from a bacterial infection of the gingival sulcus, and several clinical studies have provided evidence to implicate
Porphyromonas gingivalis
. If
P. gingivalis
is a periodontal pathogen, it would be expected to be present in most subjects with disease and rarely detected in subjects with good periodontal health. However, in most previous studies,
P. gingivalis
has not been detected in the majority of subjects with disease, and age-matched, periodontally healthy controls were not included for comparison. The purpose of the study reported here was to compare the prevalence of
P. gingivalis
in a group with periodontitis to that of a group that is periodontally healthy. A comprehensive sampling strategy and a sensitive PCR assay were used to maximize the likelihood of detection. The target sequence for
P. gingivalis
-specific amplification was the transcribed spacer region within the ribosomal operon.
P. gingivalis
was detected in only 25% (46 of 181) of the healthy subjects but was detected in 79% (103 of 130) of the periodontitis group (
P
< 0.0001). The odds ratio for being infected with
P. gingivalis
was 11.2 times greater in the periodontitis group than in the healthy group (95% confidence interval, 6.5 to 19.2). These data implicate
P. gingivalis
in the pathogenesis of periodontitis and suggest that
P. gingivalis
may not be a normal inhabitant of a periodontally healthy dentition.
Detection of Active Infection in Nonhuman Primates with Lyme Neuroborreliosis: Comparison of PCR, Culture, and a Bioassay
Authors: Pachner Andrew R. , Zhang Wei-Fen , Schaefer Henry , Schaefer Susan , O’Neill Tim 
Published: December 2019

ABSTRACT

Ideally a diagnosis of infection of the central nervous system (CNS) is made by culture of the etiologic pathogen, but
Borrelia burgdorferi
, the causative agent of Lyme neuroborreliosis (LNB), is rarely cultured from the cerebrospinal fluid (CSF). PCR and measurement of specific antibody in the CSF also have their limitations. The role of available assays for LNB has not been studied carefully in a comparative investigation. There is a need to assess the reliability of assays and to increase the ability to document active infection in the CNS. The recent development of the nonhuman primate (NHP) model of LNB allowed us to address this need in a faithful model of human LNB. In this study we compared the abilities of PCR and culture to detect the presence of spirochetes in the CSF and brain tissue of infected NHPs and related these measures of infection to the development of anti-
B. burgdorferi
antibody. We also tested a bioassay, the mouse infectivity test (MIT), in this model. Fourteen of 16 CSFs from four NHPs were positive by at least one of these techniques. Detection of spirochetes in the CSF by PCR, the MIT, and culture was inversely related to the concomitant presence of anti-
B. burgdorferi
antibody intrathecally. The performance of any particular test was associated with the strength of the host immune response. In early CNS infection, when anti-
B. burgdorferi
antibody had not yet appeared, or in immunocompromised hosts, the MIT compared favorably to culture and PCR for infected NHPs; antibody in the CSF was the most useful assay for immunocompetent NHPs.
Characterization of Monoclonal Antibodies to the 44-Kilodalton Major Outer Membrane Protein of the Human Granulocytic Ehrlichiosis Agent
Authors: Kim Hyung-Yong , Rikihisa Yasuko 
Published: December 2019

ABSTRACT

The major outer membrane proteins (OMPs) of the human granulocytic ehrlichiosis (HGE) agent, with molecular sizes of 44 to 47 kDa, are immunodominant antigens in human infection. Monoclonal antibodies (MAbs) to the OMPs were made by immunizing BALB/c mice with the purified HGE agent and then by fusing spleen cells with myeloma cells. The immunologic specificities of three MAbs (3E65, 5C11, and 5D13) were examined with five human HGE agent isolates and one tick isolate. By Western blot analysis, all three MAbs recognized the HGE agent but not
Ehrlichia chaffeensis
,
Ehrlichia sennetsu
,
Ehrlichia canis
, or their host cells. MAb 3E65 reacted with a 44-kDa protein in the homologous human isolate but not in the remaining five isolates. The two remaining MAbs recognized proteins with molecular sizes of 44 to 47 kDa in all six isolates. Western blot results with the OMP fraction of the six isolates were consistent with results with the whole HGE agent. Immunofluorescent-antibody staining and immunogold labeling with these MAbs showed that these antigens were primarily present on the membrane of the HGE agent. MAbs 5C11 and 5D13 recognized the recombinant 44-kDa protein by Western immunoblot analysis, but MAb 3E65 did not. Passive immunization with MAb 3E65 was more effective in protecting mice from HGE agent infection than with MAbs 5C11 and 5D13. These MAbs would be useful for analyzing the role of the major OMP antigens in HGE agent infection and for serodiagnosis.
Genetic Heterogeneity of Clinical Strains of
Yersinia enterocolitica
Traced by Ribotyping and Relationships between Ribotypes, Serotypes, and Biotypes
Authors: Lobato M. J. , Landeras E. , González-Hevia M. A. , Mendoza M. C. 
Published: December 2019

ABSTRACT

A series of 74
Yersinia enterocolitica
clinical strains collected in a Spanish region and 10 reference strains, assigned to nine serotypes and five biotypes, were analyzed by ribotyping procedures. Riboprobing, performed separately with
Hin
dIII and
Bgl
I and using an
rrn
operon as the probe, generated 13 and 11 ribotypes (discrimination index [DI] = 0.56 and 0.55), respectively. PCR ribotyping, performed with primers complementary to conserved regions of 16S and 23S rRNA genes, generated 13 ribotypes (DI = 0.56). A combination of data from the three procedures allowed for further discrimination into 17 combined ribotypes (DI = 0.83). The dendrogram obtained by cluster analysis of data from riboprobing indicated a high heterogeneity of the ribosomal DNA regions of the strains under study (similarities between 10 and 92%), which were grouped into three clusters at a similarity level of 0.32. The major cluster included 10 branches, and 7 of these formed a subcluster (similarity coefficient, >83%) represented by strains of serotype O:3 and biotype 2, 3, or 4. The second cluster included four branches, represented by strains belonging to seven non-O:3 serotypes, biotypes 1A and 2, and two of these branches included pyrazinamidase-positive as well as pyrazinamidase-negative strains. The remaining three branches, represented by O:3–biotype 4 strains, formed a third cluster weakly related to the others. Data from this study showed that
Y. enterocolitica
O:3 organisms assigned to a prevalent and endemic lineage and non-O:3 organisms assigned to three other less-frequent lineages are circulating and causing human disease in the Spanish region under study.
Antigenemia in Patients with Paracoccidioidomycosis: Detection of the 87-Kilodalton Determinant during and after Antifungal Therapy
Authors: Gómez B. L. , Figueroa J. I. , Hamilton A. J. , Diez S. , Rojas M. , Tobón A. M. , Hay R. J. , Restrepo A. 
Published: December 2019

ABSTRACT

Serological diagnosis and follow-up of paracoccidioidomycosis (PCM) patients have relied mainly on the detection of antibody responses by using techniques such as complement fixation (CF) and immunodiffusion. We recently described a novel inhibition enzyme-linked immunosorbent assay (inh-ELISA) which proved to be useful in the diagnosis of PCM via the detection of an 87-kDa determinant in patient sera (B. L. Gomez, J. I. Figueroa, A. J. Hamilton, B. Ortiz, M. A. Robledo, R. J. Hay, and A. Restrepo, J. Clin. Microbiol. 35:3278–3283, 1997). This test has now been assessed as a means of following up PCM patients. A total of 24 PCM patients, classified according to their clinical presentation (6 with the acute form of the disease, of whom two had AIDS, 12 with the multifocal form of the disease, and 6 with the unifocal form of the disease), were studied. The four human immunodeficiency virus-negative patients with acute PCM showed a statistically significant decrease in circulating antigen levels after the start of antifungal therapy. Antigen levels in this group became negative by our criteria (≤2.3 μg/ml) before week 20 and remained so in three of four of these patients. In contrast, the two AIDS patients who also presented with the acute form of PCM showed no statistically significant decrease in circulating antigen levels even after 68 weeks of therapy. Taken together as a group, the patients with the multifocal form showed a statistically significant decrease in antigenemia after 28 weeks of therapy. In addition, five of six patients with the unifocal form became antigen negative by week 40. Antigen level decrease mirrored clinical cure in the majority of patients in all clinical groups; in contrast, measurement of anti-PCM antibodies via the CF test showed wide fluctuations in titers during the follow-up period. The inh-ELISA for the detection of the 87-kDa
Paracoccidioides brasiliensis
determinant would appear to be a valuable additional tool in the follow-up of PCM patients.
Detection and Typing of
Borrelia burgdorferi
Sensu Lato in
Ixodes ricinus
Ticks Attached to Human Skin by PCR
Authors: Liebisch Gabriele , Sohns Bettina , Bautsch Wilfried 
Published: December 2019

ABSTRACT

Live
Ixodes ricinus
ticks attached to humans residing in Germany were examined for borreliae by dark-field microscopy and PCR.
Borrelia
species were identified by 16S rRNA sequence analysis, which showed the presence of several species, some not yet defined, and a high prevalence of multiply infected ticks.
Comparison of the QUANTIPLEX HIV-1 RNA 2.0 Assay with the AMPLICOR HIV-1 MONITOR 1.0 Assay for Quantitation of Levels of Human Immunodeficiency Virus Type 1 RNA in Plasma of Patients Receiving Stavudine-Didanosine Combination Therapy
Authors: Segondy Michel , Izopet Jacques , Pellegrin Isabelle , Montes Brigitte , Dumon Beatrice , Pasquier Christophe , Peeters Martine , Fleury Herve J. A. , Puel Jacqueline , Reynes Jacques 
Published: December 2019

ABSTRACT
We compared the QUANTIPLEX HIV-1 RNA 2.0 assay with the AMPLICOR HIV-1 MONITOR 1.0 assay for quantitation of human immunodeficiency virus type 1 (HIV-1) RNA in plasma in the Stadi trail, which evaluated a stavudine plus didanosine combination therapy in 52 patients. HIV-1 RNA baseline values measured with AMPLICOR HIV-1 MONITOR 1.0 were significantly higher than those measured with QUANTIPLEX HIV-1 RNA 2.0, and decreases in HIV-1 RNA levels from baseline were also found to be significantly higher when measured with the AMPLICOR HIV-1 MONITOR 1.0 assay. The frequency of HIV-1 RNA levels below the lower limit of quantitation was significantly higher with QUANTIPLEX HIV-1 RNA 2.0 than with AMPLICOR HIV-1 MONITOR 1.0. Reanalysis of these results by an ultrasensitive procedure of AMPLICOR HIV-1 MONITOR 1.0 or by a modified version of the test that included additional primers adapted for non-B HIV-1 clades yielded greater differences between the QUANTIPLEX HIV-1 RNA 2.0 assay and the AMPLICOR HIV-1 MONITOR 1.0 assay. Our results indicate that a valid comparison of the virological efficacies obtained with different antiretroviral drug regimens requires the use of the same viral load quantitation procedure; further standardization between the different HIV-1 RNA quantitation kits is therefore needed.
Determination of 16S rRNA Sequences of Enterococci and Application to Species Identification of Nonmotile
Enterococcus gallinarum
Isolates
Authors: Patel Robin , Piper Kerryl E. , Rouse Mark S. , Steckelberg James M. , Uhl Jim R. , Kohner Peggy , Hopkins Marlene K. , Cockerill Franklin R. , Kline Bruce C. 
Published: December 2019

ABSTRACT

The 16S rRNA sequences of enterococcal species
E. faecium
,
E. faecalis
,
E. gallinarum
,
E. casseliflavus/flavescens
,
E. dispar
,
E. pseudoavium
,
E. sulfureus
,
E. malodoratus
,
E. raffinosus
,
E. cecorum
,
E. hirae
,
E. saccharolyticus
,
E. seriolicida
,
E. mundtii
,
E. avium
,
E. durans
,
E. columbae
, and
E. solitarius
are presented herein. These data were utilized to confirm the species identification of two nonmotile
E. gallinarum
isolates which had been previously phenotypically identified as
E. faecium
. The implications of this finding are discussed.
Effect of Oxygen on Growth of
Mycobacterium ulcerans
in the BACTEC System
Authors: Palomino J. C. , Obiang A. M. , Realini L. , Meyers W. M. , Portaels F. 
Published: December 2019

ABSTRACT

The effect of low oxygen concentration on the growth of 15 strains of
Mycobacterium ulcerans
was evaluated in the BACTEC system. Reduced oxygen tension enhanced the growth of
M. ulcerans
, suggesting that this organism has a preference for microaerobic environments. Application of this observation may improve rates of isolation of
M. ulcerans
in primary culture from clinical samples and promote isolation of the bacterium from environmental sources.