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Visual detection of the human metapneumovirus using reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye.
Virol J. 2012 Jul 27; 9:138.VJ

Abstract

BACKGROUND

Human metapneumovirus (hMPV) is a major cause of acute respiratory infections ranging from wheezing to bronchiolitis and pneumonia in children worldwide. The objective of this study is to develop a visual reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of hMPV and applied to the clinical samples.

RESULTS

In this study, visual RT-LAMP assay for hMPV was performed in one step with the addition of hydroxynaphthol blue (HNB), and were used to detect respiratory samples. Six primers, including two outer primers (F3 and B3), two inner primers (FIP, BIP) and two loop primers (LF and LB), were designed for hMPV N gene by the online software. Moreover, the RT-LAMP assay showed good specificity and no cross-reactivity was observed with human rhinovirus (HRV), human respiratory syncytial Virus (RSV), or influenza virus A/PR/8/34 (H1N1). The detection limit of the RT-LAMP assay was approximately ten viral RNA copies, lower than that of traditional reverse transcriptase polymerase chain reaction (RT-PCR) 100 RNA copies. In the 176 nasopharyngeal samples, 23 (13.1%) were conformed as hMPV positive by RT-LAMP, but 18 (10.2%) positive by RT-PCR.

CONCLUSION

Compared with conventional RT-PCR, the visual hMPV RT-LAMP assay performed well in the aspect of detect time, sensitivity, specificity and visibility. It is anticipated that the RT-LAMP will be used for clinical tests in hospital or field testing during outbreaks and in emergency.

Authors+Show Affiliations

State Key Laboratory for Molecular Virology and Genetic Engineering, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Ying-Xin Road 100, Xuan Wu District, Beijing, 100052, Peoples Republic of China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22838725

Citation

Wang, Xiang, et al. "Visual Detection of the Human Metapneumovirus Using Reverse Transcription Loop-mediated Isothermal Amplification With Hydroxynaphthol Blue Dye." Virology Journal, vol. 9, 2012, p. 138.
Wang X, Zhang Q, Zhang F, et al. Visual detection of the human metapneumovirus using reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye. Virol J. 2012;9:138.
Wang, X., Zhang, Q., Zhang, F., Ma, F., Zheng, W., Zhao, Z., Bai, Y., & Zheng, L. (2012). Visual detection of the human metapneumovirus using reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye. Virology Journal, 9, 138. https://doi.org/10.1186/1743-422X-9-138
Wang X, et al. Visual Detection of the Human Metapneumovirus Using Reverse Transcription Loop-mediated Isothermal Amplification With Hydroxynaphthol Blue Dye. Virol J. 2012 Jul 27;9:138. PubMed PMID: 22838725.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Visual detection of the human metapneumovirus using reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye. AU - Wang,Xiang, AU - Zhang,Qian, AU - Zhang,Fang, AU - Ma,Fenlian, AU - Zheng,Wenzhi, AU - Zhao,Zhihui, AU - Bai,Yinglong, AU - Zheng,Lishu, Y1 - 2012/07/27/ PY - 2011/11/09/received PY - 2012/07/06/accepted PY - 2012/7/31/entrez PY - 2012/7/31/pubmed PY - 2013/3/27/medline SP - 138 EP - 138 JF - Virology journal JO - Virol. J. VL - 9 N2 - BACKGROUND: Human metapneumovirus (hMPV) is a major cause of acute respiratory infections ranging from wheezing to bronchiolitis and pneumonia in children worldwide. The objective of this study is to develop a visual reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of hMPV and applied to the clinical samples. RESULTS: In this study, visual RT-LAMP assay for hMPV was performed in one step with the addition of hydroxynaphthol blue (HNB), and were used to detect respiratory samples. Six primers, including two outer primers (F3 and B3), two inner primers (FIP, BIP) and two loop primers (LF and LB), were designed for hMPV N gene by the online software. Moreover, the RT-LAMP assay showed good specificity and no cross-reactivity was observed with human rhinovirus (HRV), human respiratory syncytial Virus (RSV), or influenza virus A/PR/8/34 (H1N1). The detection limit of the RT-LAMP assay was approximately ten viral RNA copies, lower than that of traditional reverse transcriptase polymerase chain reaction (RT-PCR) 100 RNA copies. In the 176 nasopharyngeal samples, 23 (13.1%) were conformed as hMPV positive by RT-LAMP, but 18 (10.2%) positive by RT-PCR. CONCLUSION: Compared with conventional RT-PCR, the visual hMPV RT-LAMP assay performed well in the aspect of detect time, sensitivity, specificity and visibility. It is anticipated that the RT-LAMP will be used for clinical tests in hospital or field testing during outbreaks and in emergency. SN - 1743-422X UR - https://www.unboundmedicine.com/medline/citation/22838725/Visual_detection_of_the_human_metapneumovirus_using_reverse_transcription_loop_mediated_isothermal_amplification_with_hydroxynaphthol_blue_dye_ L2 - https://virologyj.biomedcentral.com/articles/10.1186/1743-422X-9-138 DB - PRIME DP - Unbound Medicine ER -