bluuebunny/crossref_metadata_2025_split huggingface dataset:
Simplified Protocol for Pulsed-Field Gel Electrophoresis Analysis of
Streptococcus pneumoniae
Authors: McEllistrem M. Catherine , Stout Janet E. , Harrison Lee H. 
Published: May 2021

ABSTRACT

A variety of pulsed-field gel electrophoresis (PFGE) protocols for the molecular subtyping of
Streptococcus pneumoniae
have been reported; most are time-consuming and complex. We sought to modify reference PFGE protocols to reduce the time required while creating high-quality gels. Only protocol modifications that resulted in high-quality banding patterns were considered. The following protocol components were modified. Lysis enzymes (lysozyme, mutanolysin, and RNase A) were deleted in a stepwise fashion, and then the lysis buffer was deleted. Lysis and digestion were accomplished in a single step with EDTA and
N
-lauroyl sarcosine (ES; pH 8.5 to 9.3) incubation at 50°C in the absence of proteinase K. All enzymes except the restriction enzyme were omitted. A minimum incubation time of 6 h was required to achieve high-quality gels. All of the reactions were performed within 9 h, and the total protocol time from lysis to gel completion was reduced from 3 days to only 36 h. Combining lysis and digestion into a single step resulted in a substantial reduction in the time required to perform PFGE for
S. pneumoniae
. The ES solution may have caused cell lysis by activating
N
-acetylmuramyl-
l
-alanine amidase, the pneumococcal autolysin.
Evaluation of a Combination Rapid Immunoassay for Detection of
Giardia
and
Cryptosporidium
Antigens
Authors: Chan Raymond , Chen Jing , York Mary K. , Setijono Norman , Kaplan Raymond L. , Graham Fitzroy , Tanowitz Herbert B. 
Published: May 2021

ABSTRACT

A combination cassette format nonenzymatic rapid immunoassay for detection of
Giardia
and
Cryptosporidium
antigens was evaluated by using 556 patient stool specimens from three clinical laboratories. This assay (Genzyme Diagnostics
Contrast Giardia/Cryptosporidium
), which can be used with fresh or formalin-fixed specimens, had unadjusted sensitivities and specificities of 96.1 and 98.5% for
Giardia
and 100 and 98.7% for
Cryptosporidium
, respectively, in this study.
Evaluation of the BACTEC MGIT 960 and the MB/BacT Systems for Recovery of Mycobacteria from Clinical Specimens and for Species Identification by DNA AccuProbe
Authors: Alcaide Fernando , Benítez Miguel Angel , Escribà Josep M. , Martín Rogelio 
Published: May 2021

ABSTRACT

A total of 120 mycobacterial isolates were recovered from 1,068 clinical specimens. Of these, 82.5% were in MGIT 960, 83.3% were in MB/BacT, 80% were in BACTEC 460, and 70% were on Löwenstein-Jensen medium. Mean times to detection of
Mycobacterium tuberculosis
(
n
= 96) were significantly shorter with MGIT 960 (12.6 days,
P
= 0.003) and BACTEC 460 (11.8 days,
P
< 0.001) than with MB/BacT (15.9 days). Although, MGIT 960 showed the lowest rate of recovery of
M. kansasii
genotype I (64.3%), the earliest growth was detected with this system (8.9 days). Low and similar rates of contamination were obtained with MGIT 960 (3.3%) and MB/BacT (3%). The AccuProbe test for identification showed excellent sensitivities with MGIT 960 (96.8%) and MB/BacT (100%) cultures. In addition to being nonradiometric, both MGIT 960 and MB/BacT are accurate, rapid, and labor-saving detection systems which could replace the radiometric method.
Convenient Test for Screening Metallo-β-Lactamase-Producing Gram-Negative Bacteria by Using Thiol Compounds
Authors: Arakawa Yoshichika , Shibata Naohiro , Shibayama Keigo , Kurokawa Hiroshi , Yagi Tetsuya , Fujiwara Hiroshi , Goto Masafumi 
Published: May 2021

ABSTRACT

A simple disk diffusion test was constructed for detection of IMP-1-type metallo-β-lactamase-producing gram-negative bacteria. Two Kirby-Bauer disks containing ceftazidime (CAZ) and a filter disk containing a metallo-β-lactamase inhibitor were used in this test. Several IMP-1 inhibitors such as thiol compounds including 2-mercaptopropionic acid, heavy metal salts, and EDTA were evaluated for this test. Two CAZ disks were placed on a Mueller-Hinton agar plate on which a bacterial suspension was spread according to the method recommended by the National Committee for Clinical Laboratory Standards. The distance between the disks was kept to about 4 to 5 cm, and a filter disk containing a metallo-β-lactamase inhibitor was placed near one of the CAZ disks within a center-to-center distance of 1.0 to 2.5 cm. For IMP-1-producing strains, the growth-inhibitory zone between the two disks expanded, while no evident change in the shape of the growth-inhibitory zone was observed for CAZ-resistant strains producing serine β-lactamases such as AmpC or SHV-12. As a result, 2 to 3 μl of undiluted 2-mercaptopropionic acid or mercaptoacetic acid able to block IMP-1 activity gave the most reproducible and clearest results, and CAZ-resistant strains producing AmpC or extended-spectrum β-lactamases were distinguishable from IMP-1 producers by this test. A similar observation was made with IMP-1-producing clinical isolates such as
Serratia marcescens
,
Klebsiella pneumoniae
,
Escherichia coli
,
Enterobacter cloacae
,
Enterobacter aerogenes
,
Citrobacter freundii
,
Proteus vulgaris
,
Pseudomonas aeruginosa
,
Pseudomonas putida
,
Acinetobacter
spp., and
Alcaligenes xylosoxidans
. The specificity and sensitivity of this test were comparable to those of PCR analysis using
bla
IMP
-specific primers. Therefore, this convenient test would be valuable for daily use in clinical laboratories.
Quantitation of Human Immunodeficiency Virus Type 1 Group O Load in Plasma by Measuring Reverse Transcriptase Activity
Authors: Lerma J. Gerardo García , Soriano Vincent , Mas Antonio , Quiñones-Mateu Miguel E. , Arts Eric J. , Heneine Walid 
Published: May 2021

ABSTRACT
We have evaluated the use of an ultrasensitive reverse transcriptase (RT) activity assay to monitor plasma viremia in two human immunodeficiency virus type 1 (HIV-1) group O-infected patients treated with stavudine, lamivudine, and indinavir. After a initial decline in RT levels observed at 4 weeks of therapy, RT-based plasma viremia returned to baseline values at 28 or 44 weeks of treatment. The rebound in levels of RT activity was associated with the detection of phenotypic resistance to lamivudine and with the Met184Val mutation. Analysis of RT activity in plasma provides a sequence-independent means of monitoring virus loads in HIV-1 group O-infected patients.
Additional Human Papillomavirus Types Detected by the Hybrid Capture Tube Test among Samples from Women with Cytological and Colposcopical Atypia
Authors: Kónya József , Veress György , Juhász Attila , Szarka Krisztina , Sápy Tamás , Hernádi Zoltán , Gergely Lajos 
Published: May 2021

ABSTRACT

The type specificity of the human papillomavirus (HPV) Hybrid Capture Tube (HCT) test was evaluated by using typing with PCR (MY09-MY11)-restriction fragment length polymorphism (RFLP) and sequencing. All samples HCT test positive for only low-risk HPV (
n
= 15) or only high-risk HPV (
n
= 102) were confirmed, whereas 9 of 12 HCT test double-positive samples contained only high-risk HPV types as determined by PCR-RFLP. Several high-risk HPV types (HPV-53, -58, -62, -66, -CP8304, and -MM4) not included in the HCT test were indeed detected, indicating a broader detection range with retained distinction between low-risk and high-risk HPV types.
Identification of a Major Cluster of
Klebsiella pneumoniae
Isolates from Patients with Liver Abscess in Taiwan
Authors: Lau Yeu-Jun , Hu Bor-Shen , Wu Wan-Ling , Lin Yu-Hui , Chang Hwan-You , Shi Zhi-Yuan 
Published: May 2021

ABSTRACT

Klebsiella pneumoniae
has emerged as the leading liver abscess pathogen in Taiwan, with the percentage rising from 30% in the 1980s to over 80% in the 1990s. Most of the patients with
K. pneumoniae
liver abscess are diabetic and without biliary tract disease. Some patients develop serious extrahepatic complications such as endophthalmitis, meningitis, lung abscess, and necrotizing fasciitis. Pulsed-field gel electrophoresis (PFGE) was used for cluster analysis of 96 isolates from patients with liver abscess and 60 isolates from patients with other diseases. A total of 136 PFGE types were identified. Among the 96 liver abscess-associated isolates, 60 (62.5%) were classified in major cluster A. Cluster A included 41 PFGE types (types 1 to 41) which had a genetic similarity of at least 72.4% ± 9.4%. The PFGE patterns of cluster A strains are so similar that they could have originated from the same ancestor. This study demonstrates that cluster A plays an important role in the high incidence of
K. pneumoniae
liver abscess in Taiwan.
Sequencing the Gene Encoding Manganese-Dependent Superoxide Dismutase for Rapid Species Identification of Enterococci
Authors: Poyart Claire , Quesnes Gilles , Trieu-Cuot Patrick 
Published: May 2021

ABSTRACT

Simple PCR and sequencing assays that utilize a single pair of degenerate primers were used to characterize a 438-bp-long DNA fragment internal (

sodA
int

) to the
sodA
gene encoding the manganese-dependent superoxide dismutase in 19 enterococcal type strains (
Enterococcus avium
,
Enterococcus casseliflavus
,
Enterococcus cecorum
,
Enterococcus columbae
,
Enterococcus dispar
,
Enterococcus durans
,
Enterococcus faecalis
,
Enterococcus faecium
,
Enterococcus flavescens
,
Enterococcus gallinarum
,
Enterococcus hirae
,
Enterococcus malodoratus
,
Enterococcus mundtii
,
Enterococcus pseudoavium
,
Enterococcus raffinosus
,
Enterococcus saccharolyticus
,
Enterococcus seriolicida
,
Enterococcus solitarius
, and
Enterococcus sulfureus
). Sequence analysis of the

sodA
int

fragments enabled reliable identification of 18 enterococcal species, including
E. casseliflavus-E. flavescens
and
E. gallinarum
. The

sodA
int

fragments of
E. casseliflavus
and
E. flavescens
were almost identical (99.5% sequence identity), which suggests that they should be associated in a single species. Our results confirm that the
sodA
gene constitutes a more discriminative target sequence than 16S rRNA gene in differentiating closely related bacterial species.
Impact of Sample Type on Rapid Detection of Influenza Virus A by Cytospin-Enhanced Immunofluorescence and Membrane Enzyme-Linked Immunosorbent Assay
Authors: Landry Marie L. , Cohen Sandra , Ferguson David 
Published: May 2021

ABSTRACT
Cytospin-enhanced direct fluorescent-antibody assay (DFA) detected 49 (92.5%) and rapid membrane enzyme-linked immunosorbent assay (ELISA) detected 40 (75.5%) of 53 influenza virus A-positive samples. All 15 positive nasopharyngeal aspirates from children were detected by both tests. In contrast, 34 of 38 (89.5%) positive swabs from adults were detected by DFA, but only 25 (66%) were detected by ELISA.
Serological Detection of
Capillaria hepatica
by Indirect Immunofluorescence Assay
Authors: Juncker-Voss Martina , Prosl Heinrich , Lussy Helga , Enzenberg Ulrike , Auer Herbert , Nowotny Norbert 
Published: May 2021

ABSTRACT

In this paper, a serological assay for the detection of antibodies to
Capillaria hepatica
, a zoonotic parasite, is described. In the past, the only way of detecting
Capillaria hepatica
was to perform a liver biopsy. The indirect immunofluorescence (IIF) assay, based on liver sections of naturally infected mice and human serum samples, is suitable for detecting early stages of human infections and for screening purposes. No cross-reactivity with other parasitic infections was detected. We have applied the IIF assay to serum samples of 60 employees of the Zoological Garden of Vienna, Schönbrunn, Austria, and found one positive and one questionable sample.
Confirmation of
psaA
in All 90 Serotypes of
Streptococcus pneumoniae
by PCR and Potential of This Assay for Identification and Diagnosis
Authors: Morrison Katherine E. , Lake Derrick , Crook Jennifer , Carlone George M. , Ades Edwin , Facklam Richard , Sampson Jacquelyn S. 
Published: May 2021

ABSTRACT

The gene encoding the pneumococcal surface adhesin A (PsaA) protein,
psaA
, was confirmed in all
Streptococcus pneumoniae
serotypes by a newly developed PCR (
psaA
PCR) assay. Eighty-nine of the 90 serotypes amplified produced an 838-bp fragment; the exception was a serotype 16F strain acquired from the American Type Culture Collection (ATCC). Analysis of 20 additional 16F strains from the United States and Brazil showed that the gene was amplified in all 16F strains, implying that the serotype 16F ATCC strain must be a variant. The specificity of the assay was verified by the lack of signal from analysis of heterologous bacterial species (
n
= 30) and genera (
n
= 14), including viridans group streptococci. The potential of the assay for clinical application was shown by its ability to detect pneumococci in culture-positive nasopharyngeal specimens. Demonstration of
psaA
in all 90 serotypes and lack of amplification of heterologous organisms suggest that this assay could be a useful tool for detection of pneumococci and diagnosis of disease.
Development and Evaluation of a Phage Typing Scheme for
Vibrio cholerae
O139
Authors: Chakrabarti A. K. , Ghosh A. N. , Nair G. Balakrish , Niyogi S. K. , Bhattacharya S. K. , Sarkar B. L. 
Published: May 2021

ABSTRACT

The scenario of cholera that existed previously changed in 1992 and 1993 with the emergence of toxigenic
Vibrio cholerae
O139 in India. The genesis of the new serogroup formed the impetus to search for O139 phages in and around the country. A total of five newly isolated phages lytic to
V. cholerae
O139 strains were used for the development of this phage typing scheme. These phages differed from each other and also differed from the existing O1 phages in their lytic patterns, morphologies, restriction endonuclease digestion profiles, and immunological criteria. With this scheme, 500
V. cholerae
O139 strains were evaluated for their phage types, and almost all strains were found to be typeable. The strains clustered into 10 different phage types, of which type 1 (38.2%) was the dominant type, followed by type 2 (22.4%) and type 3 (18%). Additionally, a comparative study of phage types in 1993 and 1994 versus those from 1996 to 1998 for O139 strains showed a higher percentage of phage type 1 (40.5%), followed by type 3 (18.8%) during the period between 1993 and 1994, whereas phage type 2 (32.1%) was the next major type during the period from 1996 to 1998. This scheme comprising five newly isolated phages would be another useful tool in the study of the epidemiology of cholera caused by
V. cholerae
O139.
Differentiation of
Mycobacterium tuberculosis
Complex and Nontuberculous Mycobacterial Liquid Cultures by Using Peptide Nucleic Acid-Fluorescence In Situ Hybridization Probes
Authors: Drobniewski F. A. , More P. G. , Harris G. S. 
Published: May 2021

ABSTRACT

A blinded comparison of peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) with routine identification methods was performed on 74 consecutively positive mycobacterial liquid cultures. All
Mycobacterium tuberculosis
cultures (48 of 48) and 22 of 27 (81.5%) nontuberculous cultures were correctly identified (including one mixed culture). Five isolates yielded no reaction with either probe and were identified as
Mycobacterium xenopi
,
Mycobacterium fortuitum
, or
Mycobacterium flavescens
.
Quality Control Guidelines for Disk Diffusion and Broth Microdilution Antimicrobial Susceptibility Tests with Seven Drugs for Veterinary Applications
Authors: Odland Brant A. , Erwin Meredith E. , Jones Ronald N. 
Published: May 2021

ABSTRACT

This multicenter study proposes antimicrobial susceptibility (MIC and disk diffusion methods) quality control (QC) parameters for seven compounds utilized in veterinary health. Alexomycin, apramycin, tiamulin, tilmicosin, and tylosin were tested by broth microdilution against various National Committee for Clinical Laboratory Standards (NCCLS)-recommended QC organisms (
Staphylococcus aureus
ATCC 29213,
Enterococcus faecalis
ATCC 29212,
Streptococcus pneumoniae
ATCC 49619,
Escherichia coli
ATCC 25922, and
Pseudomonas aeruginosa
ATCC 27853). In addition, disk diffusion zone diameter QC limits were determined for apramycin, enrofloxacin, and premafloxacin by using
E. coli
ATCC 25922,
P. aeruginosa
ATCC 27853, and
S. aureus
ATCC 25923. The results from five or six participating laboratories produced ≥99.0% of MICs and ≥95.0% of the zone diameters within suggested guidelines. The NCCLS Subcommittee for Veterinary Antimicrobial Susceptibility Testing has recently approved these ranges for publication in the next M31 document.
Bacteremic Pneumonia Caused by a Single Clone of
Streptococcus pneumoniae
with Different Optochin Susceptibilities
Authors: Tsai Hsiu-Yuan , Hsueh Po-Ren , Teng Lee-Jene , Lee Ping-Ing , Huang Li-Min , Lee Chin-Yun , Luh Kwen-Tay 
Published: May 2021

ABSTRACT

Two isolates of
Streptococcus pneumoniae
having different optochin susceptibilities were recovered from a blood sample of a 2-year-old boy with community-acquired pneumonia. The two isolates were documented to belong to a single clone on the basis of the isolates' identical serotype (23F), antibiograms by the E-test, random amplified polymorphic DNA patterns generated by arbitrarily primed PCR, pulsed-field gel electrophoresis, and restriction fragment length polymorphism of the penicillin-binding protein genes
pbp2b
and
pbp2x
.
Polymorphism in the Pertussis Toxin Promoter Region Affecting the DNA-Based Diagnosis ofBordetellaInfection
Authors: Nygren Malin , Reizenstein Elisabet , Ronaghi Mostafa , Lundeberg Joakim 
Published: May 2021

ABSTRACTThe pertussis toxin (PT) promoter region is a frequently used target for DNA-based diagnosis of pertussis and parapertussis infections. The reported polymorphism in this region has also allowed discrimination of species in mixtures with severalBordetellaspecies by their specific PCR amplicon restriction patterns. In the present study, we investigated the degree of polymorphism in order to confirm the reliability of the assay. Five different sequence types of the amplified 239- or 249-bp region were found among the 33Bordetella pertussis,B. parapertussis, andB. bronchisepticaAmerican Type Culture Collection reference strains and patient isolates analyzed. According to the sequences that were obtained and according to the PT promoter sequences already available in the databases, restriction enzyme analysis withTaqI,BglI, andHaeII, which gave four different patterns, can be performed to reliably identifyB. pertussis,B. parapertussis, andB. bronchiseptica.
Characterization ofStreptococcus agalactiaeIsolates of Bovine and Human Origin by Randomly Amplified Polymorphic DNA Analysis
Authors: Martinez Gabriela , Harel Josee , Higgins Robert , Lacouture Sonia , Daignault Danielle , Gottschalk Marcelo 
Published: May 2021

ABSTRACTStreptococcus agalactiaeis considered one of the major causes of bovine intramammary infections. It is also found in the vaginas of women without any apparent clinical symptoms, but reports of neonatal infections, causing significant morbidity, are relatively frequent. The aim of this study was to evaluate the genetic diversity ofS. agalactiaestrains isolated from bovine milk and from asymptomatic women in Québec, Canada, by randomly amplified polymorphic DNA (RAPD) analysis. A total of 185 bovine isolates and 38 human isolates were first serotyped for capsular polysaccharide by double diffusion in agarose gel (bovine isolates) and coagglutination (human isolates). Strains were then studied by RAPD using 3 primers, designated OPS11, OPB17, and OPB18, which were selected from 12 primers. Thirty-eight percent of bovine isolates and 82% of human isolates could be serotyped. Prevalent serotypes were type III (28%) for bovine isolates and types V (26%) and III (24%) for human isolates. RAPD results showed that, taken together, all isolates (of bovine and human origin) shared 58% similarity. Ninety-four percent of these isolates were clustered in four groups (I, II, III, and IV) with 70% similarity among them. Three clusters, A (48 isolates), B (14 isolates), and C (32 isolates), with 79 to 80% similarity were identified within group IV, whereas the three other groups did not present any clusters. Despite some clustering of human isolates, relatively high diversity was seen among them. Relatively high heterogeneity was observed with the RAPD profiles, not only for field strains belonging to different serotypes but also for those within a given serotype.
Development of a TT Virus DNA Quantification System Using Real-Time Detection PCR
Authors: Kato Takanobu , Mizokami Masashi , Mukaide Motokazu , Orito Etsuro , Ohno Tomoyoshi , Nakano Tatsunori , Tanaka Yasuhito , Kato Hideaki , Sugauchi Fuminaka , Ueda Ryuzo , Hirashima Noboru , Shimamatsu Kazuhide , Kage Masayoshi , Kojiro Masamichi 
Published: May 2021

ABSTRACT
Although TT virus (TTV) was isolated from a cryptogenic posttransfusion hepatitis patient, its pathogenic role remains unclear. It has been reported that the majority of the healthy population is infected with TTV. To elucidate the differences between TTV infection in patients with liver diseases and TTV infection in the healthy population, a quantification system was developed. TTV DNA was quantified by a real-time detection PCR (RTD-PCR) assay on an ABI Prism 7700 sequence detector. With this system, TTV DNA was quantified in 78 hepatitis C virus (HCV)-infected patients (63 with elevated serum alanine aminotransferase [ALT] levels and 15 with normal ALT levels) and in 70 voluntary blood donors (BDs). The quantification range was 2.08 to 7.35 log copies/ml. The intra-assay and interassay coefficients of variation were 0.37 to 6.33% and 0.60 to 7.07%, respectively. The mean serum TTV DNA levels in the HCV-infected patients with both elevated and normal ALT levels and BDs were 3.69 ± 0.89, 3.45 ± 0.76, and 3.45 ± 0.67 log copies/ml, respectively. Comparison of the serum TTV DNA levels among the HCV-infected patients revealed that they were not related to the serum ALT and HCV core protein levels or to the histopathological score on liver biopsy. This study showed that (i) the RTD-PCR assay for the detection of TTV was accurate and had a high degree of sensitivity, (ii) the mean serum TTV DNA level was similar among HCV-infected patients, irrespective of their ALT level, and also among BDs, and (iii) a high serum TTV DNA level does not affect the serum ALT and HCV levels or liver damage in HCV-infected patients.