scholarly journals Pneumocystis Jirovecii detection and comparison of multiple diagnostic methods with quantitative real-time PCR in patients with respiratory symptoms

2020 ◽  
Vol 27 (6) ◽  
pp. 1423-1427 ◽  
Author(s):  
Mohammad Y. Alshahrani ◽  
Mohammed Alfaifi ◽  
Irfan Ahmad ◽  
Ali Gaithan Alkhathami ◽  
Abdulrahim Refdan Hakami ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Respiratory Symptoms ◽  
Diagnostic Methods ◽  
Pneumocystis Jirovecii ◽  
Quantitative Real Time Pcr

scholarly journals Confirmation of multi-parallel quantitative real-time PCR as the gold standard for detecting soil-transmitted helminths in stool

2021 ◽  
Author(s):  
Marina Papaiakovou ◽  
Nils Pilotte ◽  
Julia Dunn ◽  
David TJ Littlewood ◽  
Rubén O Cimino ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Gold Standard ◽  
Low Cost ◽  
Strongyloides Stercoralis ◽  
Diagnostic Methods ◽  
Quantitative Real Time Pcr ◽  
Soil Transmitted Helminths ◽  
Stool Samples ◽  
Kappa Agreement

AbstractDue to its simplicity and cost-effectiveness, microscopy has seen extensive field-use as the diagnostic standard for the detection of soil-transmitted helminths (STH) in stool samples. However, the sensitivity of microscopy-based detection is inadequate in reduced-transmission settings where worm burden is oftentimes low. Equally problematic, eggs of closely related species oftentimes have indistinguishable morphologies, leading to species misidentification. In light of these shortcomings, the purpose of this study was to demonstrate multi-parallel quantitative real-time PCR (qPCR) as the new “gold standard” for STH detection. Accordingly, stool samples from non-endemic participants were spiked with limited numbers of eggs or larvae (1 to 40) of five different species of STH. DNA extracts were tested using two unique multi-parallel real-time PCR-based diagnostic methods. These methods employed different target sequences (ribosomal internal transcribed spacer, or highly repetitive non-coding regions), to evaluate the detection of DNA from as little as one egg per sample. There was a statistically significant kendall correlation between egg/larvae counts and qPCR from both methods for Trichuris trichiura (0.86 and 0.872 for NHM and Baylor assays) and a strong correlation (0.602 and 0.631 for NHM and Baylor assays, respectively) for Ascaris lumbricoides. Less strong but still significant was the Kendall Tau-b value for A. duodenale (0.408 for both) and for S. stercoralis (0.483 and 0.653, respectively). In addition, using field stool samples from rural Argentina both assays had fair to moderate kappa agreement (0.329-0.454), except for Strongyloides stercoralis (0.121) that both assays had slight agreement. In spite of the small cohort of samples, both qPCR assays, targeting of two independent genomic regions, provided reproducible results and we believe that, low cost multi-parallel quantitative real-time PCR-based diagnostics should supplant microscopy as the new gold standard for stool-based detection of soil transmitted helminths in public-health and community settings.

DEVELOPMENT OF MULTIPLEX REAL-TIME PCR ASSAY FOR SIX PATHOGENS: RAPID TOOL FOR DIAGNOSIS OF BACTERIAL MENINGITIS

2019 ◽  
pp. 60-66
Author(s):  
Viet Quynh Tram Ngo ◽  
Thi Ti Na Nguyen ◽  
Hoang Bach Nguyen ◽  
Thi Tuyet Ngoc Tran ◽  
Thi Nam Lien Nguyen ◽  
...  
Keyword(s):  
Bacterial Meningitis ◽  
Real Time ◽  
Real Time Pcr ◽  
Diagnostic Methods ◽  
Type B ◽  
E Coli ◽  
Pcr Assays ◽  
Set Up ◽  
Etiological Agents ◽  
Suis Serotype

Introduction: Bacterial meningitis is an acute central nervous infection with high mortality or permanent neurological sequelae if remained undiagnosed. However, traditional diagnostic methods for bacterial meningitis pose challenge in prompt and precise identification of causative agents. Aims: The present study will therefore aim to set up in-house PCR assays for diagnosis of six pathogens causing the disease including H. influenzae type b, S. pneumoniae, N. meningitidis, S. suis serotype 2, E. coli and S. aureus. Methods: inhouse PCR assays for detecting six above-mentioned bacteria were optimized after specific pairs of primers and probes collected from the reliable literature resources and then were performed for cerebrospinal fluid (CSF) samples from patients with suspected meningitis in Hue Hospitals. Results: The set of four PCR assays was developed including a multiplex real-time PCR for S. suis serotype 2, H. influenzae type b and N. meningitides; three monoplex real-time PCRs for E. coli, S. aureus and S. pneumoniae. Application of the in-house PCRs for 116 CSF samples, the results indicated that 48 (39.7%) cases were positive with S. suis serotype 2; one case was positive with H. influenzae type b; 4 cases were positive with E. coli; pneumococcal meningitis were 19 (16.4%) cases, meningitis with S. aureus and N. meningitidis were not observed in any CSF samples in this study. Conclusion: our in-house real-time PCR assays are rapid, sensitive and specific tools for routine diagnosis to detect six mentioned above meningitis etiological agents. Key words: Bacterial meningitis, etiological agents, multiplex real-time PCR

scholarly journals KLK11 promotes the activation of mTOR and protein synthesis to facilitate cardiac hypertrophy

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yi Wang ◽  
Hongjuan Liao ◽  
Yueheng Wang ◽  
Jinlin Zhou ◽  
Feng Wang ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Angiotensin Ii ◽  
Cardiac Hypertrophy ◽  
Western Blot ◽  
Real Time ◽  
Real Time Pcr ◽  
Mtor Signaling ◽  
Heart Weight ◽  
Cardiomyocyte Hypertrophy ◽  
Quantitative Real Time Pcr

Abstract Background Cardiovascular diseases have become the leading cause of death worldwide, and cardiac hypertrophy is the core mechanism underlying cardiac defect and heart failure. However, the underlying mechanisms of cardiac hypertrophy are not fully understood. Here we investigated the roles of Kallikrein 11 (KLK11) in cardiac hypertrophy. Methods Human and mouse hypertrophic heart tissues were used to determine the expression of KLK11 with quantitative real-time PCR and western blot. Mouse cardiac hypertrophy was induced by transverse aortic constriction (TAC), and cardiomyocyte hypertrophy was induced by angiotensin II. Cardiac function was analyzed by echocardiography. The signaling pathway was analyzed by western blot. Protein synthesis was monitored by the incorporation of [3H]-leucine. Gene expression was analyzed by quantitative real-time PCR. Results The mRNA and protein levels of KLK11 were upregulated in human hypertrophic hearts. We also induced cardiac hypertrophy in mice and observed the upregulation of KLK11 in hypertrophic hearts. Our in vitro experiments demonstrated that KLK11 overexpression promoted whereas KLK11 knockdown repressed cardiomyocytes hypertrophy induced by angiotensin II, as evidenced by cardiomyocyte size and the expression of hypertrophy-related fetal genes. Besides, we knocked down KLK11 expression in mouse hearts with adeno-associated virus 9. Knockdown of KLK11 in mouse hearts inhibited TAC-induced decline in fraction shortening and ejection fraction, reduced the increase in heart weight, cardiomyocyte size, and expression of hypertrophic fetal genes. We also observed that KLK11 promoted protein synthesis, the key feature of cardiomyocyte hypertrophy, by regulating the pivotal machines S6K1 and 4EBP1. Mechanism study demonstrated that KLK11 promoted the activation of AKT-mTOR signaling to promote S6K1 and 4EBP1 pathway and protein synthesis. Repression of mTOR with rapamycin blocked the effects of KLK11 on S6K1 and 4EBP1 as well as protein synthesis. Besides, rapamycin treatment blocked the roles of KLK11 in the regulation of cardiomyocyte hypertrophy. Conclusions Our findings demonstrated that KLK11 promoted cardiomyocyte hypertrophy by activating AKT-mTOR signaling to promote protein synthesis.

Quantitative real-time PCR assay to detect transgene copy number in cotton (Gossypium hirsutum)

2008 ◽  
Vol 375 (1) ◽  
pp. 150-152 ◽  
Author(s):  
Cheng Xin Yi ◽  
Jun Zhang ◽  
Ka Man Chan ◽  
Xiao Kun Liu ◽  
Yan Hong
Keyword(s):  
Gossypium Hirsutum ◽  
Real Time ◽  
Real Time Pcr ◽  
Copy Number ◽  
Quantitative Real Time Pcr ◽  
Pcr Assay ◽  
Transgene Copy Number
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