scholarly journals Comparative Clinical Evaluation of NeoPlex RB-8 with Seeplex PneumoBacter ACE for Simultaneous Detection of Eight Respiratory Bacterial Pathogens

2019 ◽  
Vol 58 (2) ◽  
Author(s):  
Jeong Woo Kim ◽  
Seong Soo Hong ◽  
In Seop Lee ◽  
Hyun Young Chi ◽  
Soo-Ok Kim ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Legionella Pneumophila ◽  
Simultaneous Detection ◽  
Nasopharyngeal Swab ◽  
Pcr Assay ◽  
Content Type ◽  
Bordetella Parapertussis ◽  
Molecular Assays ◽  
Detection Assay ◽  
Higher Sensitivity

ABSTRACT There are several convenient and accurate molecular assays to detect respiratory bacterial infection. The NeoPlex RB-8 detection kit (NeoPlex RB-8) is a new multiplex real-time PCR assay that simultaneously detects Streptococcus pneumoniae, Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, Haemophilus influenzae, Bordetella pertussis, Bordetella parapertussis, and Moraxella catarrhalis in a single test. This study compared the clinical concordance of NeoPlex RB-8 with another method, Seeplex PneumoBacter ACE detection assay (Seeplex PB ACE), which simultaneously detects S. pneumoniae, M. pneumoniae, C. pneumoniae, L. pneumophila, H. influenzae, and B. pertussis. We tested 2,137 nasopharyngeal swab and sputum specimens using both assays. For discordant Bordetella parapertussis and M. catarrhalis specimens, we also performed bidirectional sequencing. For S. pneumoniae, M. pneumoniae, C. pneumoniae, L. pneumophila, H. influenzae, and B. pertussis, which are detected by both NeoPlex RB-8 and Seeplex PB ACE, the positive and negative agreement between the two assays ranged from 91.7 to 100% (κ = 0.918 to 1). S. pneumoniae and H. influenzae were the most discordant targets and measured with higher sensitivity and specificity by NeoPlex RB-8 than Seeplex PB ACE. For Bordetella parapertussis and M. catarrhalis, which are not detected by Seeplex PB ACE, NeoPlex RB-8 sensitivity and specificity were >99%. Overall, NeoPlex RB-8 was highly comparable to Seeplex PB ACE, but NeoPlex RB-8 was more clinically accurate, with higher throughput and more convenience.

scholarly journals Multicenter Clinical Evaluation of the Automated AriesBordetellaAssay

2018 ◽  
Vol 57 (2) ◽  
Author(s):  
Ryan F. Relich ◽  
Amy Leber ◽  
Stephen Young ◽  
Ted Schutzbank ◽  
Ronald Dunn ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Clinical Evaluation ◽  
Bordetella Pertussis ◽  
Turnaround Time ◽  
False Positives ◽  
Molecular Methods ◽  
Nasopharyngeal Swab ◽  
Limits Of Detection ◽  
Content Type ◽  
Bordetella Parapertussis

ABSTRACTMolecular methods offer superior sensitivity and specificity and reduce testing turnaround time from days to hours for detection ofBordetella pertussisandBordetella parapertussis. In this study, we evaluated the performance of the automated PCR-based AriesBordetellaAssay, which detects bothB. pertussisandB. parapertussisdirectly from nasopharyngeal swab specimens. The limits of detection (LoDs) were 1,800 CFU·ml−1forB. pertussisand 213 CFU·ml−1forB. parapertussis. The assay detected 16/18 uniqueB. pertussis/B. parapertussisstrains. Of 71 potentially cross-reacting organisms, 5 generated false positives in 1/6 replicates; none of 6 additionalBordetellaspp. were erroneously detected. Specimens were stable at 20 to 25°C for at least 10 h, at 4 to 8°C for 10 days, and at temperatures not exceeding −70°C for 6 months. Of 1,052 nasopharyngeal specimens from patients with suspected pertussis, 3.0% (n= 32) wereB. pertussispositive and 0.2% (n= 2) wereB. parapertussispositive. Combining these data with AriesBordetellaAssay data from 57 nasopharyngeal samples with previously confirmedB. pertussisorB. parapertussisdata and with data from 50 contrivedB. parapertussissamples, the proportions of positive and negative agreement of the respective Aries assays with the reference assays were 97.1% and 99.0% forB. pertussisand 100% and 99.7% forB. parapertussis. The AriesBordetellaAssay provides accurate detection and distinction ofB. pertussisandB. parapertussisinfections within 2 h. (This study has been registered at ClinicalTrials.gov under registration no. NCT02862262.)

scholarly journals Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae

2015 ◽  
Vol 53 (12) ◽  
pp. 3935-3937 ◽  
Author(s):  
Daniel Golparian ◽  
Stina Boräng ◽  
Martin Sundqvist ◽  
Magnus Unemo
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr ◽  
Molecular Detection ◽  
Analytical Sensitivity ◽  
Pcr Assay ◽  
Content Type ◽  
Dna Assay ◽  
Supplementary Test

The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection ofNeisseria gonorrhoeae.

scholarly journals A Real-Time PCR Assay for Simultaneous Detection and Differentiation of Four Common Entamoeba Species That Infect Humans

2020 ◽  
Vol 59 (1) ◽  
pp. e01986-20
Author(s):  
Ibne Karim M. Ali ◽  
Shantanu Roy
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Simultaneous Detection ◽  
Small Subunit ◽  
Clinical Samples ◽  
Pcr Assay ◽  
Content Type ◽  
Rdna Sequences ◽  
Appropriate Treatment

ABSTRACTThere are over 40 species within the genus Entamoeba, eight of which infect humans. Of these, four species (Entamoeba histolytica, E. dispar, E. moshkovskii, and E. bangladeshi) are morphologically indistinguishable from each other, and yet differentiation is important for appropriate treatment decisions. Here, we developed a hydrolysis probe-based tetraplex real-time PCR assay that can simultaneously detect and differentiate these four species in clinical samples. In this assay, multicopy small-subunit (SSU) ribosomal DNA (rDNA) sequences were used as targets. We determined that the tetraplex real-time PCR can detect amebic DNA corresponding to as little as a 0.1 trophozoite equivalent of any of these species. We also determined that this assay can detect E. histolytica DNA in the presence of 10-fold more DNA from another Entamoeba species in mixed-infection scenarios. With a panel of more than 100 well-characterized clinical samples diagnosed and confirmed using a previously published duplex real-time PCR (capable of detecting E. histolytica and E. dispar), our tetraplex real-time PCR assay demonstrated levels of sensitivity and specificity comparable with those demonstrated by the duplex real-time PCR assay. The advantage of our assay over the duplex assay is that it can specifically detect two additional Entamoeba species and can be used in conventional PCR format. This newly developed assay will allow further characterization of the epidemiology and pathogenicity of the four morphologically identical Entamoeba species, especially in low-resource settings.

scholarly journals Evaluation of Copan FecalSwab as Specimen Type for Use in Xpert C. difficile Assay

2017 ◽  
Vol 55 (10) ◽  
pp. 3123-3129 ◽  
Author(s):  
Michael J. Mashock ◽  
Matthew L. Faron ◽  
Blake W. Buchan ◽  
Nathan A. Ledeboer
Keyword(s):  
Test Performance ◽  
Pcr Assay ◽  
Content Type ◽  
Molecular Assays ◽  
Specificity And Sensitivity ◽  
Collection Device ◽  
Stool Samples ◽  
Stool Specimens ◽  
Xpert Assay ◽  
Preservation Medium

ABSTRACT Liquid-based microbiology (LBM) devices incorporating flocked swabs and preservation medium ease transport of specimens and improve specimen yield compared to traditional fiber wound swabs; however, the performance of LBM collection devices has not been evaluated in many molecular assays. It is unclear how the differences in matrix and specimen loading with an LBM device will affect test performance compared to traditional collection devices. The purpose of this study was to evaluate the performance of specimens collected in FecalSwab transport medium (Copan Diagnostics, Murrieta, CA) compared to unpreserved stool using the Cepheid Xpert C. difficile assay (Cepheid, Sunnyvale, CA). Results equivalent to unpreserved stool samples were obtained when 400 μl of FecalSwab-preserved stool was employed in the Xpert assay. The positive and negative percent agreement of specimens inoculated with FecalSwab medium ( n = 281) was 97.0% (95% confidence interval [CI], 90.9 to 96.4%) and 99.4% (95% CI, 96.4 to 99.9%), respectively, compared to reference results obtained using unpreserved stool. Throughout this study, only four discrepant results occurred when comparing preserved specimens to unpreserved stool specimens in the Xpert C. difficile PCR assay. Post discrepant analysis, using the BD MAX Cdiff assay, the specificity and sensitivity both increased to 100%. The high positive and negative percent agreements observed in this study suggest that stool preserved in FecalSwab media yields equivalent results to using unpreserved stool when tested on the Xpert C. difficile assay, allowing laboratories to adopt this liquid-based microbiology collection device.

scholarly journals Multicenter Evaluation of the ePlex Respiratory Pathogen Panel for the Detection of Viral and Bacterial Respiratory Tract Pathogens in Nasopharyngeal Swabs

2017 ◽  
Vol 56 (2) ◽  
Author(s):  
N. Esther Babady ◽  
Matthew R. England ◽  
Kristen L. Jurcic Smith ◽  
Taojun He ◽  
Dona Saumya Wijetunge ◽  
...  
Keyword(s):  
Influenza A ◽  
Simultaneous Detection ◽  
Respiratory Pathogen ◽  
Influenza B Virus ◽  
Nasopharyngeal Swab ◽  
Influenza B ◽  
Content Type ◽  
Subtype B ◽  
Percent Agreement ◽  
Virus Influenza

ABSTRACT The performance of the new ePlex Respiratory Pathogen (RP) panel (GenMark Diagnostics) for the simultaneous detection of 19 viruses (influenza A virus; influenza A H1 virus; influenza A 2009 H1 virus; influenza A H3 virus; influenza B virus; adenovirus; coronaviruses [HKU1, OC43, NL63, and 229E]; human rhinovirus/enterovirus; human metapneumovirus; parainfluenza viruses 1, 2, 3, and 4; and respiratory syncytial virus [RSV] [RSV subtype A and RSV subtype B]) and 2 bacteria (Mycoplasma pneumoniae and Chlamydia pneumoniae) was evaluated. Prospectively and retrospectively collected nasopharyngeal swab (NPS) specimens (n = 2,908) were evaluated by using the ePlex RP panel, with the bioMérieux/BioFire FilmArray Respiratory Panel (BioFire RP) as the comparator method. Discordance analysis was performed by using target-specific PCRs and bidirectional sequencing. The reproducibility of the assay was evaluated by using reproducibility panels comprised of 6 pathogens. The overall agreement between the ePlex RP and BioFire RP results was >95% for all targets. Positive percent agreement with the BioFire RP result for viruses ranged from 85.1% (95% confidence interval [CI], 80.2% to 88.9%) to 95.1% (95% CI, 89.0% to 97.9%), while negative percent agreement values ranged from 99.5% (95% CI, 99.1% to 99.7%) to 99.8% (95% CI, 99.5% to 99.9%). Additional testing of discordant targets (12%; 349/2,908) confirmed the results of ePlex RP for 38% (131/349) of samples tested. Reproducibility was 100% for all targets tested, with the exception of adenovirus, for which reproducibilities were 91.6% at low virus concentrations and 100% at moderate virus concentrations. The ePlex RP panel offers a new, rapid, and sensitive “sample-to-answer” multiplex panel for the detection of the most common viral and bacterial respiratory pathogens.

scholarly journals Method Comparison of the ImmuView L. pneumophila and L. longbeachae Urinary Antigen Test with the BinaxNOW Legionella Urinary Antigen Card for Detection of Legionella pneumophila Serogroup 1 Antigen in Urine

2019 ◽  
Vol 58 (3) ◽  
Author(s):  
Paul Badoux ◽  
Lianne Kracht-Kosten ◽  
Bjorn Herpers ◽  
Sjoerd Euser
Keyword(s):  
Sensitivity And Specificity ◽  
Legionella Pneumophila ◽  
Limit Of Detection ◽  
Method Comparison ◽  
Urine Samples ◽  
Analytical Sensitivity ◽  
Antigen Test ◽  
Urinary Antigen ◽  
Urinary Antigen Test ◽  
Content Type

ABSTRACT We compared the clinical performance of the ImmuView L. pneumophila and L. longbeachae urinary antigen test (SSI Diagnostica A/S, Hillerød, Denmark) to that of the BinaxNOW Legionella urinary antigen card (Binax; Abbott, Lake Buff, IL) using urine specimens from patients suspected of having pneumonia. In total, 100 frozen urine samples (derived from 50 Legionella cases and 50 noncases) were analyzed with both tests, as were 200 nonfrozen prospectively collected samples. For urine samples from five Legionella cases and two non-Legionella cases, analytical sensitivity (limit of detection) and repeatability were examined. The urine samples from the five Legionella cases were diluted with urine samples that tested Legionella urinary antigen negative with both tests. The analyses of the 100 frozen samples resulted in a sensitivity and specificity of both ImmuView and the BinaxNOW of 96.0% (48/50) and 100% (50/50), respectively. Of the 200 nonfrozen samples, there were three samples that showed a positive result for L. pneumophila by both tests. The analyses of reproducibility showed that for the 34 (diluted) samples that were tested at two consecutive times, 33 samples showed a consistent result for both the ImmuView and the BinaxNOW tests (Cohen’s kappa values of 0.916 and 0.928). In addition, the ImmuView test may have detected two L. longbeachae-positive urine samples, although other diagnostic tests could not confirm this. Both ImmuView and BinaxNOW showed high sensitivity and specificity for the detection of L. pneumophila serogroup 1 antigen in urine samples from clinical patients with a suspected lower respiratory tract infection.

scholarly journals Comparison of Assays for Sensitive and Reproducible Detection of Cell Culture-Infectious Cryptosporidium parvum and Cryptosporidium hominis in Drinking Water

2011 ◽  
Vol 78 (1) ◽  
pp. 156-162 ◽  
Author(s):  
Anne M. Johnson ◽  
George D. Di Giovanni ◽  
Paul A. Rochelle
Keyword(s):  
Cell Culture ◽  
Drinking Water ◽  
Cryptosporidium Parvum ◽  
False Positives ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Cryptosporidium Hominis ◽  
Content Type ◽  
Detection Assay ◽  
Pcr Targeting

ABSTRACTThis study compared the three most commonly used assays for detectingCryptosporidiumsp. infections in cell culture: immunofluorescent antibody and microscopy assay (IFA), PCR targetingCryptosporidiumsp.-specific DNA, and reverse transcriptase PCR (RT-PCR) targetingCryptosporidiumsp.-specific mRNA. Monolayers of HCT-8 cells, grown in 8-well chamber slides or 96-well plates, were inoculated with a variety of viable and inactivated oocysts to assess assay performance. All assays detected infection with low doses of flow cytometry-enumeratedCryptosporidium parvumoocysts, including infection with one oocyst and three oocysts. All methods also detected infection withCryptosporidium hominis. The RT-PCR assay, IFA, and PCR assay detected infection in 23%, 25%, and 51% of monolayers inoculated with threeC. parvumoocysts and 10%, 9%, and 16% of monolayers inoculated with one oocyst, respectively. The PCR assay was the most sensitive, but it had the highest frequency of false positives with mock-infected cells and inactivated oocysts. IFA was the only infection detection assay that did not produce false positives with mock-infected monolayers. IFA was also the only assay that detected infections in all experiments with spiked oocysts recovered from Envirochek capsules following filtration of 1,000 liters of treated water. Consequently, cell culture with IFA detection is the most appropriate method for routine and sensitive detection of infectiousCryptosporidium parvumandCryptosporidium hominisin drinking water.

scholarly journals NovelP450norGene Detection Assay Used To Characterize the Prevalence and Diversity of Soil Fungal Denitrifiers

2016 ◽  
Vol 82 (15) ◽  
pp. 4560-4569 ◽  
Author(s):  
Amy Novinscak ◽  
Claudia Goyer ◽  
Bernie J. Zebarth ◽  
David L. Burton ◽  
Martin H. Chantigny ◽  
...  
Keyword(s):  
Phylogenetic Trees ◽  
Agricultural Soils ◽  
Pcr Primers ◽  
Soil Samples ◽  
Pcr Assay ◽  
Gene Encoding ◽  
Content Type ◽  
Soil Ecosystems ◽  
Detection Assay ◽  
Denitrification Genes

ABSTRACTDenitrifying fungi produce nitrous oxide (N2O), a potent greenhouse gas, as they generally lack the ability to convert N2O to dinitrogen. Contrary to the case for bacterial denitrifiers, the prevalence and diversity of denitrifying fungi found in the environment are not well characterized. In this study, denitrifying fungi were isolated from various soil ecosystems, and novel PCR primers targeting theP450norgene, encoding the enzyme responsible for the conversion of nitric oxide to N2O, were developed, validated, and used to study the diversity of cultivable fungal denitrifiers. This PCR assay was also used to detectP450norgenes directly from environmental soil samples. Fungal denitrification capabilities were further validated using an N2O gas detection assay and a PCR assay targeting thenirKgene. A collection of 492 facultative anaerobic fungi was isolated from 15 soil ecosystems and taxonomically identified by sequencing the internal transcribed spacer sequence. Twenty-seven fungal denitrifiers belonging to 10 genera had theP450norand thenirKgenes and produced N2O from nitrite. N2O production is reported in strains not commonly known as denitrifiers, such asByssochlamys nivea,Volutella ciliata,Chloridiumspp., andTrichocladiumspp. The prevalence of fungal denitrifiers did not follow a soil ecosystem distribution; however, a higher diversity was observed in compost and agricultural soils. The phylogenetic trees constructed using partialP450norandnirKgene sequences revealed that both genes clustered taxonomically closely related strains together.IMPORTANCEA PCR assay targeting theP450norgene involved in fungal denitrification was developed and validated. The newly developedP450norprimers were used on fungal DNA extracted from a collection of fungi isolated from various soil environments and on DNA directly extracted from soil. The results indicated that approximatively 25% of all isolated fungi possessed this gene and were able to convert nitrite to N2O. All soil samples from which denitrifying fungi were isolated also tested positive for the presence ofP450nor. TheP450norgene detection assay was reliable in detecting a large diversity of fungal denitrifiers. Due to the lack of homology existing betweenP450norand bacterial denitrification genes, it is expected that this assay will become a tool of choice for studying fungal denitrifiers.

scholarly journals Semiquantitative Multiplexed Tandem PCR for Detection and Differentiation of Four Theileria orientalis Genotypes in Cattle

2014 ◽  
Vol 53 (1) ◽  
pp. 79-87 ◽  
Author(s):  
Piyumali K. Perera ◽  
Robin B. Gasser ◽  
Simon M. Firestone ◽  
Lee Smith ◽  
Florian Roeber ◽  
...  
Keyword(s):  
Genomic Dna ◽  
Simultaneous Detection ◽  
Asia Pacific ◽  
Analytical Sensitivity ◽  
Pacific Region ◽  
Asia Pacific Region ◽  
Pcr Assay ◽  
Theileria Orientalis ◽  
Content Type ◽  
Specificity And Sensitivity

Oriental theileriosis is an emerging, tick-borne disease of bovines in the Asia-Pacific region and is caused by one or more genotypes of theTheileria orientaliscomplex. This study aimed to establish and validate a multiplexed tandem PCR (MT-PCR) assay using three distinct markers (major piroplasm surface protein, 23-kDa piroplasm membrane protein, and the first internal transcribed spacer of nuclear DNA), for the simultaneous detection and semiquantification of four genotypes (Buffeli, Chitose, Ikeda, and type 5) of theT. orientaliscomplex. Analytical specificity, analytical sensitivity, and repeatability of the established MT-PCR assay were assessed in a series of experiments. Subsequently, the assay was evaluated using 200 genomic DNA samples collected from cattle from farms on which oriental theileriosis outbreaks had occurred, and 110 samples from a region where no outbreaks had been reported. The results showed the MT-PCR assay specifically and reproducibly detected the expected genotypes (i.e., genotypes Buffeli, Chitose, Ikeda, and type 5) of theT. orientaliscomplex, reliably differentiated them, and was able to detect as little as 1 fg of genomic DNA from each genotype. The diagnostic specificity and sensitivity of the MT-PCR were estimated at 94.0% and 98.8%, respectively. The MT-PCR assay established here is a practical and effective diagnostic tool for the four main genotypes ofT. orientaliscomplex in Australia and should assist studies of the epidemiology and pathophysiology of oriental theileriosis in the Asia-Pacific region.

Sign in / Sign up

Export Citation Format

Share Document