scholarly journals Establishment of a Real-time RT-PCR Assay for Avian Encephalomyelitis Virus

2016 ◽  
Vol 72 (7) ◽  
pp. 418-422
Author(s):  
Qinghong Xue ◽  
Hui Guo ◽  
Zhongze Feng ◽  
Jia Wang
Keyword(s):  
Real Time ◽  
Disease Virus ◽  
Rapid Detection ◽  
Prevention And Control ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Agar Diffusion Test ◽  
Encephalomyelitis Virus ◽  
And Control

Avian encephalomyelitis is an epidemic disease caused by avian encephalomyelitis virus (AEV), which exerts egg drop, slow growth, decreased egg hatchability, rapid tremors and ataxia of the head and neck in chicks and laying hens. The availability of a robust diagnostic assay to confirm the infection of AEV is important for its prevention and control. Thus, this study aimed to develop and validate a rapid Real-time RT-PCR assay for the detection and quantification of AEV. The authors developed and validated a TaqMan-based fluorescent real-time RT-PCR with sensitivity, specificity and reproducibility for rapid detection of EAV. This assay was then applied on 52 clinical samples from 37 suspected chickens. The minimal detection limit of this method was 20 copies. No cross-reactivity with Newcastle disease virus (NDV), avian influenza virus (AIV), Marek’s disease virus (MDV) or avian leukosis virus (ALV) was observed. The intra- and inter-assay coefficients of variation were both below 2%. Of the 52 clinical samples analyzed using this assay, 33 samples were positive for AEV, yielding an agreement rate of 91.2% with RT-PCR and 88.2% with the agar diffusion test. In conclusion, we have successfully developed and validated a sensitive Real-time RT-PCR for rapid detection of AEV, which could be used in the early diagnosis, prevention and control, and epidemiological investigation of AEV.

scholarly journals A real-time RT-PCR assay for rapid detection of coxsackievirus A10

2015 ◽  
Vol 14 (4) ◽  
pp. 17496-17504 ◽  
Author(s):  
C.Y. Mu ◽  
A.Y. Wang ◽  
C. Chen ◽  
L. Zhao ◽  
Z. Li
Keyword(s):  
Real Time ◽  
Rapid Detection ◽  
Rt Pcr ◽  
Pcr Assay

A TaqMan-MGB real-time RT-PCR assay with an internal amplification control for rapid detection of Muscovy duck reovirus

2020 ◽  
Vol 52 ◽  
pp. 101575
Author(s):  
Min Zheng ◽  
Xiuqin Chen ◽  
Shao Wang ◽  
Jingxiang Wang ◽  
Meiqing Huang ◽  
...  
Keyword(s):  
Real Time ◽  
Rapid Detection ◽  
Internal Amplification Control ◽  
Muscovy Duck ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Muscovy Duck Reovirus

Development of an EvaGreen based real-time RT-PCR assay for rapid detection, quantitation and diagnosis of goose calicivirus

2020 ◽  
Vol 49 ◽  
pp. 101489 ◽  
Author(s):  
Su Lin ◽  
Shizhong Zhang ◽  
Shao Wang ◽  
Kaichun Xie ◽  
Dandan Jiang ◽  
...  
Keyword(s):  
Real Time ◽  
Rapid Detection ◽  
Rt Pcr ◽  
Pcr Assay

scholarly journals Real-Time Reverse Transcription-Polymerase Chain Reaction Detection of Newcastle Disease Virus Using Light upon Extension Fluorogenic Primers

2007 ◽  
Vol 19 (4) ◽  
pp. 400-404 ◽  
Author(s):  
Márta Antal ◽  
Tibor Farkas ◽  
Péter Germán ◽  
Sándor Belák ◽  
István Kiss
Keyword(s):  
Newcastle Disease Virus ◽  
Real Time ◽  
Disease Virus ◽  
Newcastle Disease ◽  
Plasmid Copy Number ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Plasmid Copy ◽  
Infectious Dose ◽  
Efficient Detection

A real-time reverse transcriptase (RT)-PCR assay, applying light upon extension (LUX) fluorogenic primers, was developed for rapid and efficient detection of Newcastle disease virus (NDV). The method, which targets the fusion (F) protein gene of the viral genome, gave positive signal with all NDV isolates tested (32/32), while negative results were obtained with heterologous pathogens (35/35), including 13 avian influenza virus isolates. The detection limit of the assay was approximately 10+1.2 egg infectious dose (EID)50/0.2 ml and 10+2.2 EID50/0.2 ml for virus suspensions and spiked chicken fecal samples, respectively. As expressed in plasmid copy number, the procedure has a sensitivity of approximately 20 copies of the plasmid harboring the target gene. Due to its high specificity, sensitivity, and relative simplicity, the LUX RT-PCR assay provides a novel, rapid, and practical tool for the detection of NDV.

Rapid detection of Enterovirus and Coxsackievirus A10 by a TaqMan based duplex one-step real time RT-PCR assay

2017 ◽  
Vol 33 ◽  
pp. 8-10 ◽  
Author(s):  
Jingfang Chen ◽  
Rusheng Zhang ◽  
Xinhua Ou ◽  
Dong Yao ◽  
Zheng Huang ◽  
...  
Keyword(s):  
Real Time ◽  
Rapid Detection ◽  
Rt Pcr ◽  
Pcr Assay ◽  
One Step

scholarly journals Description and Validation of a Novel Real-Time RT-PCR Enterovirus Assay

2008 ◽  
Vol 54 (2) ◽  
pp. 406-413 ◽  
Author(s):  
Weston C Hymas ◽  
Wade K Aldous ◽  
Edward W Taggart ◽  
Jeffery B Stevenson ◽  
David R Hillyard
Keyword(s):  
Sequence Analysis ◽  
Single Nucleotide Polymorphisms ◽  
Real Time ◽  
Clinical Samples ◽  
Nucleotide Polymorphisms ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Single Nucleotide ◽  
The Real ◽  
Nontranslated Region

Abstract Background: Enteroviruses are a leading cause of aseptic meningitis in adult and pediatric populations. We describe the development of a real-time RT-PCR assay that amplifies a small target in the 5′ nontranslated region upstream of the classical Rotbart enterovirus amplicon. The assay includes an RNA internal control and incorporates modified nucleotide chemistry. Methods: We evaluated the performance characteristics of this design and performed blinded parallel testing on clinical samples, comparing the results with a commercially available RT-PCR assay (Pan-Enterovirus OligoDetect kit) that uses an enzyme immunoassay–like plate end detection. Results: We tested 778 samples and found 14 discrepant samples between the 2 assays. Of these, the real-time assay detected 6 samples that were negative by the OligoDetect kit, 5 of which were confirmed as positive by sequence analysis using an alternative primer set. Eight discrepant samples were positive by the OligoDetect kit and real-time negative, with 6 confirmed by sequencing. Overall, detection rates of 97% and 96% were obtained for the OligoDetect kit and real-time assays, respectively. Sequence analysis revealed the presence of a number of single nucleotide polymorphisms in the targeted region. The comparative sensitivities of the 2 assays were equivalent, with the limit of detection for the real-time assay determined to be approximately 430 copies per milliliter in cerebrospinal fluid. Conclusions: This novel real-time enterovirus assay is a sensitive and suitable assay for routine clinical testing. The presence of single nucleotide polymorphisms can affect real-time PCR assays.

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