TaqMan-probe-based multiplex real-time RT-qPCR for simultaneous detection of GoAstV, GPV, and GoCV

Yu, Jingquan; Zou, Junwei; Li, Xuan; Pan, Ying; Mu, Yuanyuan; Li, Shuyan; Wang, Juhua; Xu, Fei; Wang, Yong

doi:10.1016/j.psj.2022.102396

Cited by 2 publications

(2 citation statements)

References 27 publications

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“…According to the results of clinical sample detection, an increasing tendency towards mixed infection caused by AMDV, MEV and CDV was found, suggesting that a detection method capable of detecting and distinguishing AMDV, MEV and CDV simultaneously is a prerequisite for the epidemiological investigation of mink infections. The multiplex real-time PCR (qPCR) method decreases detection costs, enhances e ciency and enhances accuracy, and these advantages give qPCR a primary position in disease screening and clinical settings [19][20][21].…”

Section: Discussionmentioning

confidence: 99%

Multiplex one-step RT‒qPCR assays for simultaneous detection of AMDV, MEV and CDV

Cao,

Xu,

Zhao

et al. 2024

Preprint

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Background: Aleutian mink disease, mink viral enteritis and canine distemper are known as the three most serious diseases that cause great economic loss in the mink industry. In clinical practice, aleutian mink disease virus (AMDV), mink enteritis virus (MEV) and canine distemper virus (CDV) are common mixed infections, and they have similar clinical symptoms, such as diarrhoea. Therefore, a rapid and accurate differential diagnosis method for use on mink ranches is essential for the control of these three pathogens. Here, we developed multiplex one-step real-time quantitative PCR (RT‒qPCR) assays for the simultaneous detection and quantification of AMDV, MEV and CDV by using three primers and probes based on the conserved NS1, VP2 and N genes, respectively. Results: The results showed that the established method was less likely to cross-react with other mink pathogens, with a detection sensitivity of 25 copies/μL and a coefficient of variation less than 3.51%. Moreover, the interference experiment showed that the presence of AMDV, MEV and CDV templates at different concentrations would not interfere with the detection results. Furthermore, two hundred clinical samples of mink with diarrhoea were simultaneously analysed using multiplex RT‒qPCR and single RT‒qPCR, the Kappa values were all greater than 0.921, indicating that there was a high degree of coincidence between the two detection methods. Conclusions: In conclusion, multiplex RT‒qPCR exhibited high speciﬁcity, sensitivity, and reproducibility, indicating that this method can be used as a reliable and speciﬁc tool for the differential detection and quantiﬁcation of AMDV, MEV and CDV.

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Section: Discussionmentioning

confidence: 99%

Multiplex one-step RT‒qPCR assays for simultaneous detection of AMDV, MEV and CDV

Cao,

Xu,

Zhao

et al. 2024

Preprint

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“…At present, the following detection methods have been reported: real-time reverse transcription polymerase chain reaction (RT-PCR) [27][28][29][30], TaqMan-probe-based real-time RT-qPCR [31,32], peptide-based ELISA [33], indirect competitive ELISA [34], one-step reverse transcription loop-mediated isothermal amplification [35], reverse transcriptionenzymatic recombinase amplification coupled with a CRISPR-Cas12a system [14], and immunochromatographic strip assay [36]. In the past few years, ddPCR has undergone rapid development and has been widely applied for the detection and quantitative analysis of a variety of viruses [37,38].…”

Section: Discussionmentioning

confidence: 99%

The Development of a Sensitive Droplet Digital Polymerase Chain Reaction Test for Quantitative Detection of Goose Astrovirus

Shi,

Jin,

Zhang

et al. 2024

Viruses

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(1) Goose astrovirus (GAstV) is a novel emerging pathogen that causes significant economic losses in waterfowl farming. A convenient, sensitive, and specific detection method for GAstV in field samples is important in order to effectively control GAstV. Droplet digital polymerase chain reaction (ddPCR) is a novel, sensitive, good-precision, and absolute quantitation PCR technology which does not require calibration curves. (2) In this study, we developed a ddPCR system for the sensitive and accurate quantification of GAstV using the conserved region of the ORF2 gene. (3) The detection limit of ddPCR was 10 copies/µL, ~28 times greater sensitivity than quantitative real-time PCR (qPCR). The specificity of the test was determined by the failure of amplification of other avian viruses. Both ddPCR and qPCR tests showed good repeatability and linearity, and the established ddPCR method had high sensitivity and good specificity to GAstV. Clinical sample test results showed that the positive rate of ddPCR (88.89%) was higher than that of qPCR (58.33%). (4) As a result, our results suggest that the newly developed ddPCR method might offer improved analytical sensitivity and specificity in its GAstV measurements. The ddPCR could be widely applied in clinical tests for GAstV infections.

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TaqMan-probe-based multiplex real-time RT-qPCR for simultaneous detection of GoAstV, GPV, and GoCV

Cited by 2 publications

References 27 publications

Multiplex one-step RT‒qPCR assays for simultaneous detection of AMDV, MEV and CDV

Multiplex one-step RT‒qPCR assays for simultaneous detection of AMDV, MEV and CDV

The Development of a Sensitive Droplet Digital Polymerase Chain Reaction Test for Quantitative Detection of Goose Astrovirus

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