Development and evaluation of a quantitative polymerase chain reaction for aquatic Streptococcus agalactiae based on the groEL gene

2020 ◽  
Vol 129 (1) ◽  
pp. 63-74
Author(s):  
W.J. Leigh ◽  
R.N. Zadoks ◽  
J.Z. Costa ◽  
A. Jaglarz ◽  
K.D. Thompson
Keyword(s):  
Polymerase Chain Reaction ◽  
Streptococcus Agalactiae ◽  
Quantitative Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Groel Gene ◽  
Polymerase Chain

scholarly journals Use of a new multiplex quantitative polymerase chain reaction based assay for simultaneous detection of Neisseria meningitidis, Escherichia coli K , Streptococcus agalactiae, and Streptococcus pneumoniae

2021 ◽  
Author(s):  
Nastaran Hemmati ◽  
Farhad Nikkhahi ◽  
Amir Javadi ◽  
Sahar Eskandarion ◽  
Seyed Mahmoud Amin Marashi
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction ◽  
Streptococcus Pneumoniae ◽  
Bacterial Meningitis ◽  
Neisseria Meningitidis ◽  
Streptococcus Agalactiae ◽  
Predictive Value ◽  
Quantitative Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Polymerase Chain

Background and Objectives: Neisseria meningitidis, Escherichia coli K , Streptococcus agalactiae, and Streptococcus pneumoniae cause 90% of bacterial meningitis. Almost all infected people die or have irreversible neurological complica- tions. Therefore, it is essential to have a diagnostic kit with the ability to quickly detect these fatal infections. Materials and Methods: The project involved 212 patients from whom cerebrospinal fluid samples were obtained. After total genome extraction and performing multiplex quantitative polymerase chain reaction (qPCR), the presence or absence of each infectious factor was determined by comparing with standard strains. Results: The specificity, sensitivity, positive predictive value, and negative predictive value calculated were 100%, 92.9%, 50%, and 100%, respectively. So, due to the high specificity and sensitivity of the designed primers, they can be used instead of bacterial culture that takes at least 24 to 48 hours. Conclusion: The remarkable benefit of this method is associated with the speed (up to 3 hours) at which the procedure could be completed. It is also worth noting that this method can reduce the personnel unintentional errors which may occur in the laboratory. On the other hand, as this method simultaneously identifies four common factors that cause bacterial meningitis, it could be used as an auxiliary method diagnostic technique in laboratories particularly in cases of emergency medicine.

AN IMPROVED METHOD FOR INHIBITOR FREE NUCLEIC ACIDS FROM POLYPHENOL RICH LEAVES OF MUSA SPP

2017 ◽  
Vol 23 (1) ◽  
Author(s):  
N.NANDHA KUMAR ◽  
K. SOURIANATHA SUNDARAM ◽  
D. SUDHAKAR ◽  
K.K. KUMAR
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Proteinase K ◽  
Chain Reaction ◽  
Banana Plant ◽  
High Molecular Weight Dna ◽  
Musa Spp ◽  
Leaf Tissues ◽  
Polymerase Chain

Excessive presence of polysaccharides, polyphenol and secondary metabolites in banana plant affects the quality of DNA and it leads to difficult in isolating good quality of DNA. An optimized modified CTAB protocol for the isolation of high quality and quantity of DNA obtained from banana leaf tissues has been developed. In this protocol a slight increased salt (NaCl) concentration (2.0M) was used in the extraction buffer. Polyvinylpyrrolidone (PVP) and Octanol were used for the removal of polyphenols and polymerase chain reaction (PCR) inhibitors. Proteins like various enzymes were degraded by Proteinase K and removed by centrifugation from plant extract during the isolation process resulting in pure genomic DNA, ready to use in downstream applications including PCR, quantitative polymerase chain reaction (qPCR), ligation, restriction and sequencing. This protocol yielded a high molecular weight DNA isolated from polyphenols rich leaves of Musa spp which was free from contamination and colour. The average yields of total DNA from leaf ranged from 917.4 to 1860.9 ng/ìL. This modified CTAB protocol reported here is less time consuming 4-5h, reproducible and can be used for a broad spectrum of plant species which have polyphenol and polysaccharide compounds.

scholarly journals Screening of Additive Formulations Enables Off-Chip Drop Reverse Transcription Quantitative Polymerase Chain Reaction of Single Influenza A Virus Genomes

2021 ◽  
Vol 93 (10) ◽  
pp. 4365-4373
Author(s):  
Emma Kate Loveday ◽  
Geoffrey K. Zath ◽  
Dimitri A. Bikos ◽  
Zackary J. Jay ◽  
Connie B. Chang
Keyword(s):  
Polymerase Chain Reaction ◽  
Influenza A Virus ◽  
Reverse Transcription ◽  
Influenza A ◽  
Quantitative Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Virus Genomes

scholarly journals Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Yang Zhang ◽  
Chunyang Dai ◽  
Huiyan Wang ◽  
Yong Gao ◽  
Tuantuan Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Highly Sensitive ◽  
One Step ◽  
Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

scholarly journals Accuracy of quantitative polymerase chain reaction in samples of frozen and paraffin-embedded healthy skin for the diagnosis of canine visceral leishmaniasis

2017 ◽  
Vol 69 (6) ◽  
pp. 1443-1450 ◽  
Author(s):  
M.P. Campos ◽  
M.F. Madeira ◽  
D.A. Silva ◽  
M.S. Solcà ◽  
O.M. Espíndola ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Healthy Skin ◽  
Sample Collection ◽  
Chain Reaction ◽  
Dna Recovery ◽  
Ffpe Samples ◽  
Fresh Frozen ◽  
Polymerase Chain ◽  
Commercial Kits

ABSTRACT The purpose of the present work was to evaluate the accuracy of quantitative polymerase chain reaction (qPCR) performed on samples of fresh frozen tissue (FT) and formalin-fixed, paraffin-embedded (FFPE) healthy skin. This is a validation study conducted with samples from 46 dogs from an endemic area in Brazil. After sample collection, DNA extractions were conducted using commercial kits and qPCR was oriented to kinetoplast DNA (kDNA) targets of the Leishmania infantum species. The results obtained for the FFPE samples showed 63.6% sensitivity and 77.1% specificity, whereas those obtained for the FT samples showed 100% and 48.6%, respectively. Poor agreement was observed for the results of the qPCR technique with FT and FFPE samples. Our results suggest freezing as the most suitable conservation method for the formation of sample databases considering DNA recovery

scholarly journals Rapid diagnosis of invasive pulmonary aspergillosis by quantitative polymerase chain reaction using bronchial lavage fluid

2002 ◽  
Vol 72 (1) ◽  
pp. 27-30 ◽  
Author(s):  
Masahito Kawazu ◽  
Yoshinobu Kanda ◽  
Susumu Goyama ◽  
Masataka Takeshita ◽  
Yasuhito Nannya ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Invasive Pulmonary Aspergillosis ◽  
Lavage Fluid ◽  
Rapid Diagnosis ◽  
Chain Reaction ◽  
Pulmonary Aspergillosis ◽  
Bronchial Lavage ◽  
Polymerase Chain
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