scholarly journals A Retrospective Study about the Impact of Switching from Nested PCR to Multiplex Real-Time PCR on the Distribution of the Human Papillomavirus (HPV) Genotypes

Medicina ◽  
2019 ◽  
Vol 55 (8) ◽  
pp. 418 ◽  
Author(s):  
Prete ◽  
Ronga ◽  
Addati ◽  
Magrone ◽  
Abbasciano ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
Real Time ◽  
Real Time Pcr ◽  
Nested Pcr ◽  
Diagnostic Methods ◽  
Pcr Assay ◽  
Hpv Prevalence ◽  
Significant Difference ◽  
Hpv Genotypes ◽  
The Impact

Background and objectives: Human papillomavirus (HPV) is the most prevalent etiological agent of viral sexually-transmitted infection. This study retrospectively evaluated the impact of a switch to a real-time PCR assay in the HPV prevalence and genotypes distribution by a quasi-experimental before-and-after approach. Materials and Methods: In total, 1742 samples collected from 1433 patients were analyzed at the UOC Microbiology and Virology of Policlinico of Bari, Italy. HPV DNA detection was performed using initially nested PCR and subsequently multiplex real-time PCR assay. Results: Statistically significant difference in HPV overall prevalence after the introduction of the real-time assay was not detected (48.97% vs. 50.62%). According to different extraction-DNA amplification methods, differences were observed in the prevalence rates of HPV-45, 68, 40, 42, and 43. The lowest prevalence for HPV-45 was observed in the Magna Pure-Real Time PCR group, while HPV-68, 40, 42, and 43 were less observed in the Qiagen-Real Time PCR group. After, a multivariate logistic regression, an increase in the prevalence of HPV-42 (aOR: 4.08, 95% CI: 1.71–9.73) was associated with the multiplex real-time PCR assay. Conclusions: Although this study is a not a direct comparison between two diagnostic methods because it has a sequential structure, it serves to verify the impact of a new molecular assay on HPV distribution. Moreover, the stability of HPV prevalence over time suggests that the population composition and the behavioral variables did not likely change during the observation period. Our study proposes that the introduction of a molecular test for HPV detection may be related to changes of HPV genotypes distribution.

scholarly journals The Sensitivity of the Real-time PCR and Nested-PCR for Detection of Coxiella burnetii in Milk Samples

2017 ◽  
Vol 4 (2) ◽  
pp. 11
Author(s):  
Mojtaba Bonyadian ◽  
Hamdollah Moshtaghi ◽  
Hamidreza Kazemeini
Keyword(s):  
Real Time ◽  
Coxiella Burnetii ◽  
Real Time Pcr ◽  
Nested Pcr ◽  
Pcr Assay ◽  
Milk Samples ◽  
Bulk Milk ◽  
Significant Difference ◽  
Pcr Assays ◽  
Nested Pcr Assays

Coxiella burnetii is the causative agent of Q-fever, a widespread zoonosis. In domestic animals infection remains either asymptomatic or presents as infertility or abortion. Clinical presentation in humans can range from mild flu-like illness to acute pneumonia and hepatitis. In humans serology is the gold standard for diagnosis but is inadequate for early case detection, so real-time PCR and nested-PCR assays were developed in this study to measure amounts of C.burnetii shed in milk. Our study was to assess the sensitivity of the realtime PCR and nested-PCR for detection of Coxiella burnetii in bovine bulk milk samples from dairy herds in 3 provinces (Chaharmahal and Bakhtiari , Isfahan and Yazd) of Iran. In the present study, 300 bulk milk samples from 89 dairy cattle herds were tested for C. burnetii using real-time PCR and nested-PCR assays. The animals which their milk samples collected for this study were clinically healthy. In total, 74 of 300 (24.7%) cow milk samples were positive in real-time PCR assay and 26 of 300 (8.7%) samples were positive in nested-PCR assay. McNemar test shows a significant difference in detection of C. burnetii between real-time PCR and nested-PCR. Also the results of this study indicate those clinically healthy dairy cows are important sources of C. burnetii infection in Iran.

Roles of the multiplex real-time PCR assay and β-D-glucan in a high-risk population for intra-abdominal candidiasis (IAC)

2019 ◽  
Vol 58 (6) ◽  
pp. 789-796 ◽  
Author(s):  
J Fortún ◽  
M J Buitrago ◽  
F Gioia ◽  
E Gómez-Gª de la Pedrosa ◽  
M E Alvarez ◽  
...  
Keyword(s):  
High Risk ◽  
Real Time ◽  
Real Time Pcr ◽  
Volume Overload ◽  
Diagnostic Methods ◽  
Practice Volume ◽  
High Risk Population ◽  
Pcr Assay ◽  
European Consensus ◽  
Abdominal Candidiasis

Abstract Multiplex quantitative real-time PCR (MRT-PCR) using blood can improve the diagnosis of intra-abdominal candidiasis (IAC). We prospectively studied 39 patients with suspected IAC in the absence of previous antifungal therapy. Blood cultures, MRT-PCR, and β-D-glucan (BDG) in serum were performed in all patients. IAC was defined according to the 2013 European Consensus criteria. For MRT-PCR, the probes targeted the ITS1 or ITS2 regions of ribosomal DNA. Candidaemia was confirmed only in four patients (10%), and IAC criteria were present in 17 patients (43.6%). The sensitivity of MRT-PCR was 25% but increased to 63.6% (P = .06) in plasma obtained prior to volume overload and transfusion; specificity was above 85% in all cases. BDG performance was improved using a cutoff > 260 pg/ml, and improvement was not observed in samples obtained before transfusion. In this cohort of high risk of IAC and low rate of bloodstream infection, the performance of non-culture-based methods (MRT-PCR or BDG) was moderate but may be a complementary tool given the limitations of diagnostic methods available in clinical practice. Volume overload requirements, in combination with other factors, decrease the accuracy of MRT-PCR in patients with IAC.

scholarly journals Clinical Validation of the HPV-Risk Assay, a Novel Real-Time PCR Assay for Detection of High-Risk Human Papillomavirus DNA by Targeting the E7 Region

2014 ◽  
Vol 52 (3) ◽  
pp. 890-896 ◽  
Author(s):  
A. T. Hesselink ◽  
J. Berkhof ◽  
M. L. van der Salm ◽  
A. P. van Splunter ◽  
T. H. Geelen ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
High Risk ◽  
Real Time ◽  
Real Time Pcr ◽  
Clinical Validation ◽  
Pcr Assay ◽  
Papillomavirus Dna

Demonstration of human papillomavirus type 16 positivity by real time-PCR assay in patients with colorectal carcinoma

2015 ◽  
Vol 70 ◽  
pp. S75
Author(s):  
Ayca Ozer Durmuslu ◽  
Guumllendam Bozdayi ◽  
Aylin Altay ◽  
Ozgur Ekinci ◽  
Ayse Dursun ◽  
...  
Keyword(s):  
Human Papillomavirus ◽  
Colorectal Carcinoma ◽  
Real Time ◽  
Real Time Pcr ◽  
Pcr Assay ◽  
Human Papillomavirus Type 16

scholarly journals Evaluation of the ELITe InGenius PCR Platform for Detection of Mycoplasma pneumoniae

2019 ◽  
Vol 57 (6) ◽  
Author(s):  
Arthur H. Totten ◽  
Sixto M. Leal ◽  
Amy E. Ratliff ◽  
Li Xiao ◽  
Donna M. Crabb ◽  
...  
Keyword(s):  
Real Time ◽  
Mycoplasma Pneumoniae ◽  
Real Time Pcr ◽  
Growth Media ◽  
Test Accuracy ◽  
Analytical Sensitivity ◽  
Pcr Assay ◽  
Predictive Values ◽  
Content Type ◽  
Significant Difference

ABSTRACT Mycoplasma pneumoniae is the leading cause of bacterial community-acquired pneumonia in persons of all ages. Due to the fastidious nature of this bacterium and the necessary specialized growth media, nucleic acid amplification testing is currently the most reliable means for patient diagnostics. Analytical sensitivity, specificity, reproducibility, and clinical performance of the ELITe InGenius automated PCR platform with its MGB Alert M. pneumoniae real-time PCR research use only reagents (ELITechGroup, Inc., Bothell, WA) were compared with those of a laboratory-developed real-time PCR assay targeting repMp1 for detection of M. pneumoniae. The ELITe InGenius PCR assay successfully detected 31 distinct M. pneumoniae clinical isolates and reference strains, and there was no cross-reactivity with other mollicutes, Gram-positive bacteria, or Gram-negative bacteria. In testing 223 clinical samples, the ELITe InGenius PCR showed 95.79% and 99.22% positive and negative agreement with the repMp1 assay, respectively. Additionally, the ELITech platform showed 98.91% positive and 96.95% negative predictive values, and there was no significant difference detected between the two assays (McNemar’s test, P = 0.375). The ELITe InGenius PCR assay limit of detection was 0.16 CFU/PCR test or 4.16 genome copies (GCs)/test. Accuracy, instrument ease-of-use, and decreased hands-on time make the ELITe InGenius platform suitable for detection of M. pneumoniae directly from clinical specimens.

scholarly journals Evaluation of the Multiplex Real-Time PCR DermaGenius® Assay for the Detection of Dermatophytes in Hair Samples from Senegal

2021 ◽  
Vol 8 (1) ◽  
pp. 11
Author(s):  
Mouhamadou Ndiaye ◽  
Rosalie Sacheli ◽  
Khadim Diongue ◽  
Caroline Adjetey ◽  
Rajae Darfouf ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Melting Curve ◽  
Diagnostic Methods ◽  
Melting Curve Analysis ◽  
Microsporum Canis ◽  
Molecular Tools ◽  
Pcr Assay ◽  
Epidermophyton Floccosum ◽  
Hair Samples

For the successful treatment of dermatophytoses, especially tinea capitis, there is a need for accurate and rapid diagnostic methods. A lot of recent literature has focused on the detection of dermatophytes directly on sample material such as nails, hair and skin scrapings. Molecular tools offer the ability to rapidly diagnose dermatophytosis within 48 h. This study aimed to compare the results of a commercial real-time PCR (real-time PCR) assay DermaGenius®(DG) 2.0 complete multiplex kit with those of conventional diagnostic methods (direct microscopy and culture). A total of 129 hair samples were collected in Dakar (Senegal) from patients suspected of dermatophytosis. DG was applied for the molecular detection of Candida albicans, Trichophyton rubrum/soudanense, T. interdigitale, T. tonsurans, T. mentagrophytes, T. violaceum, Microsporum canis, M. audouinii, Epidermophyton floccosum, T. benhamiae and T. verrucosum. Dermatophytes species and C. albicans were differentiated by melting curve analysis. The sensitivity and specificity of the PCR assay were 89.3% and 75.3%, respectively. DG PCR was significantly more sensitive than culture (p < 0.001). DG PCR is fast and robust to contamination. In this paper, the main questions discussed were the replacement of culture by a broad-spectrum fungal real-time PCR and the implementation of DG PCR into a routine laboratory in Senegal.

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