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
Received: June 24, 2013 Revised: September 26, 2013 Accepted: September 29, 2013
Corresponding author. Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hua 1st Road, Kweishan, Taoyuan 333, Taiwan, ROC.
Background and purpose:
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.