bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genotypic Variation and Stability of Four Variable-Number Tandem Repeats and Their Suitability for Discriminating Strains ofMycobacterium leprae
Authors: Truman Richard , Fontes Amanda B. , de Miranda Antonio B. , Suffys Philip , Gillis Thomas 
Published: June 2004

ABSTRACTIt has not been possible to distinguish different strains ofMycobacterium lepraeaccording to their genetic sequence. However, the genome contains several variable-number tandem repeats (VNTR), which have been used effectively in strain typing of other bacteria. To determine their suitability for differentiatingM. leprae, we developed PCR systems to amplify 5 different VNTR loci and examined a battery of 12M. lepraestrains derived from patients in different regions of the United States, Brazil, Mexico, and the Philippines, as well as from wild armadillos and a sooty mangabey monkey. We found diversity at four VNTR (D = 0.74), but one system (C16G8) failed to yield reproducible results. Alleles for the GAA VNTR varied in length from 10 to 16 copies, those for AT17varied in length from 10 to 15 copies, those for GTA varied in length from 9 to 12 copies, and those for TA18varied in length from 13 to 20 copies. Relatively little variation was seen with interspecies transfer of bacilli or during short-term passage of strains in nude mice or armadillos. The TA18locus was more polymorphic than other VNTR, and genotypic variation was more common after long-term expansion in armadillos. Most strain genotypes remained fairly stable in passage, but strain Thai-53 showed remarkable variability. Statistical cluster analysis segregated strains and passage samples appropriately but did not reveal any particular genotype associable with different regions or hosts of origin. VNTR polymorphisms can be used effectively to discriminateM. lepraestrains. Inclusion of additional loci and other elements will likely lead to a robust typing system that can be used in community-based epidemiological studies and select clinical applications.
Evaluation and Comparison of Two Commercial Enzyme-Linked Immunosorbent Assay Kits for Detection of Antigenically Diverse Human Noroviruses in Stool Samples
Authors: Burton-MacLeod Jonathan A. , Kane Erin M. , Beard Rachel S. , Hadley Leslie A. , Glass Roger I. , Ando Tamie 
Published: June 2004

ABSTRACTTwo recently commercialized enzyme-linked immunosorbent assay kits, the SRSV(II)-AD (Denka Seiken Co. Ltd., Tokyo, Japan) and IDEIA NLV (DakoCytomation Ltd., Ely, United Kingdom) kits, that detect human norovirus (HuNV) antigens in stool samples were evaluated to assess whether they could be used instead of reverse transcription-PCR (RT-PCR) for routine diagnosis. The sensitivities and specificities of the two kits were tested with a panel of 103 stool samples containing HuNVs of 4 and 10 genetic subgroups within genogroups I and II (GI and GII), respectively, and 39 stool samples containing other enteric viruses. The Denka kit had a high sensitivity (>70% for 10 of the 14 subgroups) but a specificity of only 69%, and the Dako kit had a low sensitivity (<30% for 6 GII subgroups) but a high specificity of 100%. Statistical analysis suggests that HuNVs of four subgroups (subgroups GII/2, GII/5, GII/6, and GII/n) are likely to elude detection by the Dako kit. The two kits also demonstrated differences in reactivities. While the Dako kit discriminated between the GI and GII antigens of HuNVs, the Denka kit cross-reacted with samples containing all GI and GII subgroups of HuNVs. Moreover, the Denka kit also reacted with samples containing human sapovirus (HuSV). We demonstrate that the cross-reactivity of the Denka kit is not due to specific reactions with HuNV and HuSV antigens. These results indicate that neither the Denka kit nor the Dako kit has all the performance characteristics required to replace the RT-PCR methods used to detect HuNVs.
Variability of theChlamydia trachomatis omp1Gene Detected in Samples from Men Tested in Male-Only Saunas in Melbourne, Australia
Authors: Lister Nichole A , Tabrizi Sepehr N. , Fairley Christopher K. , Smith Anthony , Janssen Peter H. , Garland Suzanne 
Published: June 2004

ABSTRACTA recent screening program in Melbourne, Victoria, Australia, has shown thatChlamydia trachomatisis an important infection among men who frequent male-only saunas. To evaluate theC. trachomatisisolates circulating in local saunas, theC. trachomatis-positive samples collected during the program underwent amplification and sequencing of theomp1gene, and the corresponding serovars were deduced. Forty-sevenC. trachomatis-positive samples collected (from October 2001 to September 2002) from 39 men were evaluated. The deduced serovars found, in descending order of prevalence, were D, G, and J; and serovars B, E, F, and H were each found in single samples. The seven different serovars identified in the study sample indicate that local saunas are a reservoir of multipleC. trachomatisstrains, possibly maintained by the introduction of new patrons or regular patrons who have been exposed toC. trachomatiselsewhere. No significant genetic variants were found, as most variable positions were silent and were detected only in single samples.
Rapid and Specific Detection of
Escherichia coli
Clonal Group A by Gene-Specific PCR
Authors: Johnson James R. , Owens Krista , Manges Amee R. , Riley Lee W. 
Published: June 2004

ABSTRACT

PCR primers specific for the recently described antimicrobial resistance-associated
Escherichia coli
clonal group A (CGA), a widespread cause of drug-resistant urinary tract infections in the United States, were devised on the basis of a novel single-nucleotide polymorphism identified within the housekeeping gene
fumC
, i.e., C288T. In comparison with two reference PCR-based fingerprinting methods, ERIC2 PCR and random amplified polymorphic DNA (RAPD) analysis, a PCR assay incorporating the new primers provided 100% sensitivity and 100% specificity for the detection of CGA among 138 diverse clinical and reference
E. coli
isolates.
E. coli
reference (ECOR) strain 47 was shown to be a member or a close relative of CGA (by ERIC2 PCR and RAPD analysis, respectively) and yielded a positive assay result. The new CGA-specific PCR assay, which exhibited interlaboratory reproducibility and stability under various experimental conditions, should allow the rapid and specific detection of CGA by any laboratory equipped for diagnostic PCR.
Comparison of the Roche LightCycler
vanA
/
vanB
Detection Assay and Culture for Detection of Vancomycin-Resistant Enterococci from Perianal Swabs
Authors: Sloan L. M. , Uhl J. R. , Vetter E. A. , Schleck C. D. , Harmsen W. S. , Manahan J. , Thompson R. L. , Rosenblatt J. E. , Cockerill F. R. 
Published: June 2004

ABSTRACT

We compared the performance characteristics of a real-time PCR method, the LightCycler
vanA
/
vanB
detection assay (Roche Diagnostics Corporation, Indianapolis, Ind.) to that of Enterococcosel agar (BBL, Sparks, Md.) for direct detection of vancomycin-resistant enterococci (VRE) from 894 perianal stool swabs. For 421 of 894 swabs, the result for LightCycler PCR was compared to an Enterococcosel plate containing vancomycin at 6 μg/ml; for the remaining 473 swabs, the result for LightCycler PCR was compared to an Enterococcosel plate containing 8 μg/ml vancomycin. The LightCycler method produced considerably more positive results than either the Enterococcosel plate containing vancomycin at 6 μg/ml (
n
= 25 versus
n
= 11; sensitivity, 100%; specificity, 97%; positive predictive value [PPV], 42%; negative predictive value [NPV], 100%) or the Enterococcosel plate containing vancomycin at 8 μg/ml (
n
= 31 versus
n
= 10; sensitivity, 100%; specificity, 95%; PPV, 32%; NPV, 100%). When possible, additional testing, including culture, LightCycler PCR, and/or a conventional PCR method (PCR-restriction fragment length polymorphism assay), were performed on either the original specimens or original cultures or subsequent specimens for cases in which the original specimen was positive by LightCycler PCR but the Enterococcosel plate was negative. This additional testing demonstrated positive results for 7 of 14 (50%) evaluable discordant specimens which initially tested as LightCycler PCR positive but culture negative using the Enterococcosel plate containing vancomycin at 6 μg/ml and 12 of 17 (71%) evaluable discordant specimens which initially tested as LightCycler positive but culture negative using the Enterococcosel plate containing vancomycin at (8 μg/ml). These results demonstrate that the LightCycler VRE detection assay is considerably more sensitive than the standard culture method for detecting VRE directly from perianal swab specimens. The LightCycler assay also provides results much faster than culture (∼3.5 versus ≥72 h). The use of this test could have important implications for the effective control and prevention of nosocomial outbreaks of VRE.
Real-Time PCR Assay Using Fine-Needle Aspirates and Tissue Biopsy Specimens for Rapid Diagnosis of Mycobacterial Lymphadenitis in Children
Authors: Bruijnesteijn van Coppenraet E. S. , Lindeboom J. A. , Prins J. M. , Peeters M. F. , Claas E. C. J. , Kuijper E. J. 
Published: June 2004

ABSTRACT

A real-time PCR assay was developed to diagnose and identify the causative agents of suspected mycobacterial lymphadenitis. Primers and probes for the real-time PCR were designed on the basis of the internal transcribed spacer sequence, enabling the recognition of the genus
Mycobacterium
and the species
Mycobacterium avium
and
M
.
tuberculosis
. The detection limit for the assay was established at 1,100 CFU/ml of pus, and the specificity tests showed no false-positive reaction with other mycobacterial species and other pathogens causing lymphadenitis. From 67 children with suspected mycobacterial lymphadenitis based on a positive mycobacterial skin test, 102 samples (58 fine-needle aspirates [FNA] and 44 tissue specimens) were obtained. The real-time PCR assay detected a mycobacterial infection in 48 patients (71.6%), whereas auramine staining and culturing were positive for 31 (46.3%) and 28 (41.8%) of the patients. The addition of the real-time PCR assay to conventional diagnostic tests resulted in the recognition of 13 more patients with mycobacterial disease. These results indicate that the real-time PCR is more sensitive than conventional staining and culturing techniques (
P
= 0.006). The
M
.
avium
-specific real-time PCR was positive for 38 patients, and the
M
.
tuberculosis
-specific real-time PCR was positive for 1 patient. Analysis of 27 patients from whom FNA and tissue biopsy specimens were collected revealed significantly more positive real-time PCR results for FNA than for tissue biopsy specimens (
P
= 0.003). Samples from an age-matched control group of 50 patients with PCR-proven cat scratch disease were all found to be negative by the real-time PCR. We conclude that this real-time PCR assay with a sensitivity of 72% for patients with lymphadenitis and a specificity of 100% for the detection of atypical mycobacteria can provide excellent support for clinical decision making in children with lymphadenitis.
Identification of Clinically Relevant Viridans Group Streptococci by Sequence Analysis of the 16S-23S Ribosomal DNA Spacer Region
Authors: Chen Chao Chien , Teng Lee Jene , Chang Tsung Chain 
Published: June 2004

ABSTRACT

The feasibility of sequence analysis of the 16S-23S ribosomal DNA (rDNA) intergenic spacer (ITS) for the identification of clinically relevant viridans group streptococci (VS) was evaluated. The ITS regions of 29 reference strains (11 species) of VS were amplified by PCR and sequenced. These 11 species were
Streptococcus anginosus
,
S. constellatus
,
S. gordonii
,
S. intermedius
,
S. mitis
,
S. mutans
,
S. oralis
,
S. parasanguinis
,
S. salivarius
,
S. sanguinis
, and
S. uberis
. The ITS lengths (246 to 391 bp) and sequences were highly conserved among strains within a species. The intraspecies similarity scores for the ITS sequences ranged from 0.98 to 1.0, except for the score for
S. gordonii
strains. The interspecies similarity scores for the ITS sequences varied from 0.31 to 0.93. Phylogenetic analysis of the ITS regions revealed that evolution of the regions of some species of VS is not parallel to that of the 16S rRNA genes. One hundred six clinical isolates of VS were identified by the Rapid ID 32 STREP system (bioMérieux Vitek, Marcy l'Etoile, France) and by ITS sequencing, and the level of disagreement between the two methods was 18% (19 isolates). Most isolates producing discrepant results could be unambiguously assigned to a specific species by their ITS sequences. The accuracy of using ITS sequencing for identification of VS was verified by 16S rDNA sequencing for all strains except strains of
S. oralis
and
S. mitis
, which were difficult to differentiate by their 16S rDNA sequences. In conclusion, identification of species of VS by ITS sequencing is reliable and could be used as an alternative accurate method for identification of VS.
Comparative Evaluation of Three Serological Methods for Detection of Human Herpesvirus 8-Specific Antibodies in Canadian Allogeneic Stem Cell Transplant Recipients
Authors: Sergerie Yan , Abed Yacine , Roy Jean , Boivin Guy 
Published: June 2004

ABSTRACT
Human herpesvirus 8 (HHV-8) has been associated with all types of Kaposi's sarcoma (KS), including posttransplantation KS. However, little is known regarding HHV-8 infections in hematopoietic stem cell transplant (SCT) recipients. In this study, we used a variety of serological assays, including in-house-developed enzyme immunoassays (EIAs) utilizing synthetic peptides corresponding to lytic viral antigens (ORFs 65 and K8.1) and a commercial EIA kit based on a whole virus lysate (Advanced Biotechnologies Inc.), as well as latent- and lytyc-antigen-based immunofluorescence assays (IFAs) to determine the seroprevalence of HHV-8 in 42 allogeneic SCT recipients from Canada. Using the two peptide-based EIA methods as for screening, HHV-8-specific antibodies were detected in five (12%) patients between days 21 and 91, although only one (2%) subject was positive for HHV-8-specific antibodies before transplantation. All positive results from these five patients were confirmed by at least one of the IFAs, with an additional patient showing seropositivity before transplantation. However, the commercial EIA was negative at all time points (days −7, 21, and 91) in those five patients. The episodes of seroconversion or reactivation were not associated with sustained viremia, since HHV-8 DNA was not detected by real-time PCR in the corresponding leukocytes and plasma of the seropositive patients. No clinical or laboratory abnormalities were clearly associated with HHV-8 seropositivity. This study confirms the utility of simple peptide-based EIA methods to assess the presence of HHV-8-specific antibodies in immunocompromised patients and emphasizes the need of conducting prospective studies to determine the source of HHV-8 infection in SCT recipients.
Prevalence of
Helicobacter pylori vacA
and
cagA
Genotypes in Ethiopian Dyspeptic Patients
Authors: Asrat Daniel , Nilsson Ingrid , Mengistu Yohannes , Kassa Endale , Ashenafi Senait , Ayenew Kiros , Wadström Torkel , Abu-Al-Soud Waleed 
Published: June 2004

ABSTRACT

A total of 300 gastric biopsy samples and 50
Helicobacter pylori
isolates were collected from Ethiopian adult dyspeptic patients. The
vacA
and
cagA
genes were detected in 90 and 79% of biopsy specimens, respectively, and in 100 and 87% of clinical isolates, respectively. Both genes were detected in 84% of the gastric biopsy samples and in 87% of the clinical isolates. Among
vacA
genotypes, the s1/m1 genotype was the most common in gastric biopsy samples (48%). The
vacA
and
cagA
positive
H. pylori
strains were detected to a higher degree in patients with chronic active gastritis (71%) than patients with other histopathological findings (29%) (
P
< 0.05).
Evaluation of a High-Throughput Repetitive-Sequence-Based PCR System for DNA Fingerprinting of
Mycobacterium tuberculosis
and
Mycobacterium avium
Complex Strains
Authors: Cangelosi Gerard A. , Freeman Robert J. , Lewis Kaeryn N. , Livingston-Rosanoff Devon , Shah Ketan S. , Milan Sparrow Joy , Goldberg Stefan V. 
Published: June 2004

ABSTRACT

Repetitive-sequence-based PCR (rep-PCR) is useful for generating DNA fingerprints of diverse bacterial and fungal species. Rep-PCR amplicon fingerprints represent genomic segments lying between repetitive sequences. A commercial system that electrophoretically separates rep-PCR amplicons on microfluidic chips, and provides computer-generated readouts of results has been adapted for use with
Mycobacterium
species. The ability of this system to type
M. tuberculosis
and
M. avium
complex (MAC) isolates was evaluated.
M. tuberculosis
strains (
n
= 56) were typed by spoligotyping with rep-PCR as a high-resolution adjunct. Results were compared with those generated by a standard approach of spoligotyping with IS
6110
-targeted restriction fragment length polymorphism (IS
6110
-RFLP) as the high-resolution adjunct. The sample included 11 epidemiologically and genotypically linked outbreak isolates and a population-based sample of 45 isolates from recent immigrants to Seattle, Wash., from the African Horn countries of Somalia, Eritrea, and Ethiopia. Twenty isolates exhibited unique spoligotypes and were not analyzed further. Of the 36 outbreak and African Horn isolates with nonunique spoligotypes, 23 fell into four clusters identified by IS
6110
-RFLP and rep-PCR, with 97% concordance observed between the two methods. Both approaches revealed extensive strain heterogeneity within the African Horn sample, consistent with a predominant pattern of reactivation of latent infections in this immigrant population. Rep-PCR exhibited 89% concordance with IS
1245
-RFLP typing of 28
M. avium
subspecies
avium
strains. For
M. tuberculosis
as well as
M. avium
subspecies
avium
, the discriminative power of rep-PCR equaled or exceeded that of RFLP. Rep-PCR also generated DNA fingerprints from
M. intracellulare
(
n
= 8) and MAC
x
(
n
= 2) strains. It shows promise as a fast, unified method for high-throughput genotypic fingerprinting of multiple
Mycobacterium
species.
Detection and Prevalence of Active Drug Efflux Mechanism in Various Multidrug-Resistant
Klebsiella pneumoniae
Strains from Turkey
Authors: Hasdemir Ufuk Over , Chevalier Jacqueline , Nordmann Patrice , Pagès Jean-Marie 
Published: June 2004

ABSTRACT

The prevalence of active drug efflux pump and porin alterations was investigated in Turkish nosocomial strains of
Klebsiella pneumoniae
exhibiting a multidrug-resistant phenotype. MICs of various antibiotics, including quinolones, chloramphenicol, tetracycline, and β-lactams, for those strains were determined either with or without the efflux pump inhibitor phenylalanine arginine β-naphthylamide (PAβN). Thirty-nine percent of the strains exhibited a PAβN-modulated resistance for quinolones, chloramphenicol, and tetracycline. In these strains, a significant increase of chloramphenicol accumulation was gained in the presence of the efflux pump inhibitor PAβN or with the energy uncoupler carbonyl cyanide
m
-chlorophenylhydrazone. Moreover, high-level expression of the membrane fusion protein AcrA, which was immunodetected in most of those isolates, suggests that the AcrAB/TolC efflux machinery contributed to their antibiotic resistance. Studies of
K. pneumoniae
porins indicated that the majority of the strains, including extended-spectrum β-lactamase producers and efflux-positive ones, presented an alteration in their sorbitol-sensitive porin (OmpK35) expression. This is the first report showing the prominent role of active drug efflux in the antibiotic resistance of nosocomial
K. pneumoniae
strains from Turkey.
Prevalence of and Risk Factors for FelineTritrichomonas foetusandGiardiaInfection
Authors: Gookin Jody L. , Stebbins Martha E. , Hunt Emily , Burlone Karen , Fulton Merritt , Hochel Robin , Talaat Miriam , Poore Matthew , Levy Michael G. 
Published: June 2004

ABSTRACTData were gathered for 117 cats from 89 catteries at an international cat show to examine prevalence and risk factors for felineTritrichomonas foetusandGiardiainfection. Prevalence ofT. foetuswas 31% among cats (36 out of 117) and catteries (28 out of 89) based on results of fecal smear examination (5 out of 36), fecal culture in modified Diamond's medium (9 out of 36), fecal culture in In Pouch TF medium (20 out of 36), or PCR amplification of the ribosomal RNA gene from feces withT. foetus-specific primers (34 out of 36). Catteries in whichT. foetuswas identified were more likely to have had a recent history of diarrhea, historical diagnosis of coccidia infection in adult cats, and a decreased number of square feet of facility per cat. Evidence did not exist for the ongoing transmission ofT. foetusby water, food, or contact with other species.
Performance of the Celera Diagnostics ViroSeq HIV-1 Genotyping System for Sequence-Based Analysis of Diverse Human Immunodeficiency Virus Type 1 Strains
Authors: Eshleman Susan H. , Hackett John , Swanson Priscilla , Cunningham Shawn P. , Drews Birgit , Brennan Catherine , Devare Sushil G. , Zekeng Léopold , Kaptué Lazare , Marlowe Natalia 
Published: June 2004

ABSTRACT

The Celera Diagnostics ViroSeq HIV-1 Genotyping System is a Food and Drug Administration-cleared, integrated system for sequence-based analysis of drug resistance mutations in subtype B human immunodeficiency virus type 1 (HIV-1) protease and reverse transcriptase (RT). We evaluated the performance of this system for the analysis of diverse HIV-1 strains. Plasma samples were obtained from 126 individuals from Uganda, Cameroon, South Africa, Argentina, Brazil, and Thailand with viral loads ranging from 2.92 to >6.0 log
10
copies/ml. HIV-1 genotyping was performed with the ViroSeq system. HIV-1 subtyping was performed by using phylogenetic methods. PCR products suitable for sequencing were obtained for 125 (99%) of the 126 samples. Genotypes including protease (amino acids 1 to 99) and RT (amino acids 1 to 321) were obtained for 124 (98%) of the samples. Full bidirectional sequence data were obtained for 95 of those samples. The sequences were categorized into the following subtypes: A1/A2 (16 samples), B (12 samples), C (13 samples), D (11 samples), CRF01_AE (9 samples), F/F2 (9 samples), G (7 samples), CRF02_AG (32 samples), H (1 sample), and intersubtype recombinant (14 samples). The performances of the individual sequencing primers were examined. Genotyping of duplicate samples in a second laboratory was successful for 124 of the 126 samples. The identity level for the sequence data from two laboratories ranged from 98 to 100% (median, 99.8%). The ViroSeq system performs well for the analysis of plasma samples with diverse non-B subtypes. The availability of this genotyping system should facilitate studies of HIV-1 drug resistance in non-subtype B strains of HIV-1.