Development of a reverse transcription recombinase polymerase amplification assay for rapid detection of human respiratory syncytial virus

Xi, Yun; Xu, Changzhi; Xie, Zhizhi; Zhu, Dingliang; Dong, Jianwen; Xiao, Gang

doi:10.1016/j.mcp.2019.03.005

Cited by 13 publications

(10 citation statements)

References 44 publications

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“…This study established a highly sensitive and specific diagnostic approach for the detection of SFTSV nucleic acids in clinical serum specimens. The detection limit of SFTSV RT-RPA assay was statistically estimated as 241 copies of target RNA per reaction, which is consistent with other published literature [19][20][21]. While the sensitivity of this RT-RPA assay is lower than conventional RT-qPCR for SFTSV, the detection limit is still far below the viremia level in most clinical SFTS patients [18].…”

Section: Discussionsupporting

confidence: 87%

“…The RNA extraction method used in this study to process serum sample still require much effort and heavy centrifuges, which might hamper the practicality of this assay in point-of-care settings. Magnetic bead-based methods [21] or one-step RPA microfluidics [31] could be employed to simplify the sample preparation process. In detection of clinical samples, the RT-RPA assay has a certain probability to produce false-negative results [32].…”

Section: Discussionmentioning

confidence: 99%

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Development and evaluation of a rapid detection assay for severe fever with thrombocytopenia syndrome virus based on reverse-transcription recombinase polymerase amplification

Zhou

Wang

Zhu

et al. 2020

Molecular and Cellular Probes

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Development and evaluation of a rapid detection assay for severe fever with thrombocytopenia syndrome virus based on reverse-transcription recombinase polymerase amplification, Molecular and Cellular Probes (2020), doi: https://doi.Table of Abbreviations: dT-FAM Thymidine nucleotide carrying FAM fluorophore dT-BHQ1 Thymidine nucleotide carrying BHQ-1 quencher HHV Human herpesvirus LAMP loop-mediated isothermal amplification LOD Limit of detection PIV Parainfluenza virus RPA Recombinase polymerase amplification RT-LAMP Reverse-transcription loop-mediated isothermal amplification RT-PCR Reverse-transcription polymerase chain reaction RT-qPCR Real-time reverse-transcription polymerase chain reaction RT-RAA Reverse-transcription recombinase-aided amplification RT-RPA Real-time reverse-transcription recombinase polymerase amplification SFTS Severe fever with thrombocytopenia syndrome SFTSV Severe fever with thrombocytopenia syndrome virus SSB Single-stranded DNA binding Abstract 1 Rapid detection of severe fever with thrombocytopenia syndrome virus (SFTSV) is 2 crucial for its control and surveillance. In this study, a rapid isothermal real-time 3 reverse-transcription recombinase polymerase amplification (RT-RPA) assay was 4 developed for the detection of SFTSV. The detection limit at 95% probability was 241 5 copies per reaction. A test of 120 serum samples of suspected severe fever with 6 thrombocytopenia syndrome (SFTS) patients revealed that the sensitivity and 7 specificity of the RT-RPA assay was approximately 96.00% (95%CI: 80.46%-99.79%) 8 and 98.95% (95% CI: 94.28%-99.95%), respectively; the kappa value was 0.9495 (P 9 ＜0.001). The Bland-Altman analysis showed that 87.50% of the different data points 10 were located within the 95% limits of agreement, indicating a good correlation 11 between the results from RT-RPA assays and those of RT-qPCR assays. In conclusion, 12 the rapid and efficient RT-RPA assay can be a promising candidate for point-of-care 13 detection method of SFTSV. 14 15 Keywords: Severe fever with thrombocytopenia syndrome virus; Recombinase 16 polymerase amplification; Detection assay Severe fever with thrombocytopenia syndrome virus (SFTSV), belonging to the 19 family Phenuiviridae, the genus Banyangvirus, the species Huaiyangshan 20 banyangvirus, was first identified as the causative pathogen of the outbreak of severe 21 fever with thrombocytopenia syndrome (SFTS) in the central and northeast regions of 22 China in 2009 [1]. During the following decade, this emerging endemic tick-borne 23 disease has been reported in many countries worldwide, including South Korea, Japan, 24 United Arab Emirates, and the United States [2-5]. Delayed admission due to 25 nonspecific clinical manifestations, along with rapid progression to multiple organ 26 dysfunction in severe cases led to a high case fatality rate of 6.40~30% in SFTS 27 patients [6, 7]. 28 Currently, detection methods of SFTSV comprise nucleic acid testing, serological 29 detection, and virus isolation [1, 8, 9]. Due to the long turnaround...

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Section: Discussionsupporting

confidence: 87%

Section: Discussionmentioning

confidence: 99%

Development and evaluation of a rapid detection assay for severe fever with thrombocytopenia syndrome virus based on reverse-transcription recombinase polymerase amplification

Zhou

Wang

Zhu

et al. 2020

Molecular and Cellular Probes

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“…HCoV-NL63 was even detected with 1 copy/reaction. This sensitivity of this method outperforms that of other detection methods [21][22][23][24][25].…”

Section: Discussionmentioning

confidence: 81%

Simultaneous Detection of Seven Human Coronaviruses by Multiplex PCR and MALDI-TOF MS

Kang

et al. 2021

COVID

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Human coronaviruses (HCoVs) are associated with a range of respiratory symptoms. The discovery of severe acute respiratory syndrome (SARS)-CoV, Middle East respiratory syndrome, and SARS-CoV-2 pose a significant threat to human health. In this study, we developed a method (HCoV-MS) that combines multiplex PCR with matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), to detect and differentiate seven HCoVs simultaneously. The HCoV-MS method had high specificity and sensitivity, with a 1–5 copies/reaction detection limit. To validate the HCoV-MS method, we tested 163 clinical samples, and the results showed good concordance with real-time PCR. Additionally, the detection sensitivity of HCoV-MS and real-time PCR was comparable. The HCoV-MS method is a sensitive assay, requiring only 1 μL of a sample. Moreover, it is a high-throughput method, allowing 384 samples to be processed simultaneously in 30 min. We propose that this method be used to complement real-time PCR for large-scale screening studies.

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“…A reverse transcription recombinase polymerase amplification assay (RT-RPA) is nucleic acid probe based on novel isothermal amplification technique which has been widely employed to detect human RSV. The results exemplified that concurrence rates between RT-RPA assay and qRT-PCR assay for clinical samples was 96%, demonstrating that RT-RPA assay holds better diagnostic presentation on clinical samples in remote rural areas in developing countries (Xi et al 2019). Using quantitative PCR (qPCR) for common respiratory viruses and for two genes (CCL8/CXCL11) is recognized to be extremely upregulated in viral infections.…”

Section: Pcr/rt-pcr-based Immunodetectionmentioning

confidence: 87%

Advanced Immunotechnological Methods for Detection and Diagnosis of Viral Infections: Current Applications and Future Challenges

Bramhachari

Sheela

Prathyusha

et al. 2020

Dynamics of Immune Activation in Viral Diseases

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Diagnosis and identification of viruses is an important component of diagnostic virology laboratory. Although various modes of diagnostic methods are now available at disposal, a vast majority of the diseases across the globe remain undiagnosed. This is largely due to the overlapping undifferentiated set of symptoms across myriad set of RNA and DNA viral diseases. As such, it becomes critical to take into consideration several factors for viral diagnosis ranging from the type and quality of specimen collected, time of specimen collection, mode of transport, accuracy, specificity, sensitivity, and the type of diagnostic method used. This chapter broadly emphasizes various methods on diagnostic virology ranging from the classical methods of diagnosis to the most recently developed molecular methods of detection of virus.

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Development of a reverse transcription recombinase polymerase amplification assay for rapid detection of human respiratory syncytial virus

Cited by 13 publications

References 44 publications

Development and evaluation of a rapid detection assay for severe fever with thrombocytopenia syndrome virus based on reverse-transcription recombinase polymerase amplification

Development and evaluation of a rapid detection assay for severe fever with thrombocytopenia syndrome virus based on reverse-transcription recombinase polymerase amplification

Simultaneous Detection of Seven Human Coronaviruses by Multiplex PCR and MALDI-TOF MS

Advanced Immunotechnological Methods for Detection and Diagnosis of Viral Infections: Current Applications and Future Challenges

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