Pyrosequencing reveals an oseltamivir-resistant marker in the quasispecies of avian influenza A (H7N9) virus

Chee-Keng Mok; Shih-Cheng Chang; Guang-Wu Chen; Yu-Lun Lo; Shu-Jen Chen; Ho-Sheng Wu; Ming-Tsan Liu; Feng-Yee Chang; Tzou-Yien Lin; Shin-Ru Shih

doi:10.1016/j.jmii.2013.09.010

Pyrosequencing reveals an oseltamivir-resistant marker in the quasispecies of avian influenza A (H7N9) virus

J Microbiol Immunol Infect. 2015 Aug;48(4):465-9. doi: 10.1016/j.jmii.2013.09.010. Epub 2013 Dec 30.

Authors

Chee-Keng Mok¹, Shih-Cheng Chang², Guang-Wu Chen³, Yu-Lun Lo², Shu-Jen Chen⁴, Ho-Sheng Wu⁵, Ming-Tsan Liu⁵, Feng-Yee Chang⁵, Tzou-Yien Lin⁶, Shin-Ru Shih⁷

Affiliations

¹ Research Center for Emerging Viral Infections, Chang Gung University and Clinical Virology Laboratory, Chang Gung Memorial Hospital, Taiwan Centers for Disease Control Collaborating Laboratory of Virology, Taoyuan, Taiwan, ROC; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan, ROC.
² Research Center for Emerging Viral Infections, Chang Gung University and Clinical Virology Laboratory, Chang Gung Memorial Hospital, Taiwan Centers for Disease Control Collaborating Laboratory of Virology, Taoyuan, Taiwan, ROC.
³ Research Center for Emerging Viral Infections, Chang Gung University and Clinical Virology Laboratory, Chang Gung Memorial Hospital, Taiwan Centers for Disease Control Collaborating Laboratory of Virology, Taoyuan, Taiwan, ROC; Department of Computer Science and Information Engineering, College of Engineering, Chang Gung University, Taoyuan, Taiwan, ROC.
⁴ Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan, ROC; Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan, ROC.
⁵ Research and Diagnostics Center, Centers for Disease Control, Taipei, Taiwan, ROC.
⁶ Ministry of Health and Welfare, Executive Yuan, Taipei, Taiwan, ROC.
⁷ Research Center for Emerging Viral Infections, Chang Gung University and Clinical Virology Laboratory, Chang Gung Memorial Hospital, Taiwan Centers for Disease Control Collaborating Laboratory of Virology, Taoyuan, Taiwan, ROC; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan, ROC. Electronic address: srshih@mail.cgu.edu.tw.

PMID: 24388586
DOI: 10.1016/j.jmii.2013.09.010

Abstract

Prompt diagnosis of an oseltamivir-resistant marker is important for patient management, in particular to prevent the spread of resistant strains in the recent human H7N9 outbreak. We tailored a pyrosequencing assay to reveal neuraminidase R292K, a resistant marker found in one isolate from China, and demonstrated its performance in both sensitivity and specificity. In addition, a semi-nested polymerase chain reaction was applied, which enhanced the detection rate by at least 10-fold. We validated this assay by examining the marker in Taiwan's first imported human case and found R and K in quasispecies.

Keywords: Avian influenza A virus; H7N9; Oseltamivir-resistant marker; Pyrosequencing.

Publication types

Research Support, Non-U.S. Gov't
Validation Study

MeSH terms

Antiviral Agents / pharmacology*
Drug Resistance, Viral*
Humans
Influenza A Virus, H7N9 Subtype / drug effects*
Influenza A Virus, H7N9 Subtype / enzymology*
Influenza A Virus, H7N9 Subtype / isolation & purification
Influenza, Human / virology
Molecular Sequence Data
Mutation, Missense
Neuraminidase / genetics*
Oseltamivir / pharmacology*
RNA, Viral / genetics
Reverse Transcriptase Polymerase Chain Reaction / methods*
Sequence Analysis, DNA
Taiwan
Viral Proteins / genetics*

Substances

Antiviral Agents
RNA, Viral
Viral Proteins
Oseltamivir
NA protein, influenza A virus
Neuraminidase