Molecular detection of human H7N9 influenza A virus causing outbreaks in China

Abstract

Background: A novel subtype of influenza A virus (H7N9) was recently identified in humans. The virus is a reassortant of avian viruses, but these human isolates contain mutations [hemagglutinin (HA) Q226L and PB2 E627K] that might make it easier for the virus to adapt to mammalian hosts. Molecular tests for rapid detection of this virus are urgently needed.

Methods: We developed a 1-step quantitative real-time reverse-transcription PCR assay to detect the novel human H7N9 virus. The primer set was specific to the hemagglutinin (HA) gene of the H7N9 viruses currently causing the outbreak in China and had mismatches to all previously known avian or mammalian H7 HA sequences. In addition, the assay was evaluated using influenza A viruses of various genetic backgrounds and other negative controls.

Results: The detection limit of the assay was approximately 0.04 TCID50 (median tissue culture infective dose) per reaction. The assay specificity was high and all negative control samples, including 8 H7 viruses not closely related to the human H7N9 virus, tested negative.

Conclusions: The established assay allows rapid detection of the novel human H7N9 virus, thereby allowing better pandemic preparedness.