Comparative Evaluation of Three Preprocessing Methods for Extraction and Detection of Influenza A Virus Nucleic Acids from Sputum

Yu, Fei; Qiu, Ting; Zeng, Ying; Wang, Yiyin; Zheng, Shufa; Chen, Xiao; Chen, Yu

Human infection with avian influenza A (H7N9) has been remaining persistent in China over the recent years. For laboratory diagnosis of H7N9 virus infection, related studies and guidelines have confirmed that the nucleic acid-positive rate of lower respiratory specimens, such as sputum, airway aspirates, and bronchoalveolar lavage fluid, is higher than that of upper respiratory specimens (1, 2) . Clinicians give preference to sputum specimen for influenza virus test and genotyping because this kind of specimen is easy to obtain. However, real-time reverse transcription PCR (rRT-PCR) of sputum specimens yields false-negative results owing to the difficulty of extracting RNA from sputum containing mucus (3) . Meanwhile, viscous sputum specimens usually cannot undergo automated extraction, and thus, a pre-homogenization process is desirable before isolating nucleic acids for rRT-PCR. In fact, some influenza A virus (IAV) detection kits lack homogenization reagents, and some clinical laboratories may have applied improper preprocessing methods, such as adding normal saline (NS) and mixing by vortex.