Received on ,
Accepted on ,

Brief Communications

Molecular Detection of a Novel Human Influenza (H1N1) of Pandemic Potential by Conventional and Real-Time Quantitative RT-PCR Assays

¹ Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China
² Department of Microbiology, Queen Mary Hospital, Hong Kong SAR, China
³ Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China, and Department of Microbiology, Queen Mary Hospital, Hong Kong SAR, China
⁴ Department of Microbiology, The University of Hong Kong, Hong Kong SAR, China, and HKU-Pasteur Research Centre, Hong Kong SAR, China

^* To whom correspondence should be addressed. E-mail: llmpoon{at}hkucc.hku.hk.

BACKGROUND: Influenza A viruses are medically important viral pathogens causing significant mortality and morbidity throughout the world. The recent emergence of a novel human influenza A virus (H1N1) poses a serious health threat. Development of molecular tests for rapid detection of this virus is urgently needed.

METHODS: We developed a conventional 1-step RT-PCR assay and a 1-step quantitative real-time RTPCR assay to detect the novel H1N1 virus, but not the seasonal H1N1 viruses. An additional real-time RT-PCR that can discriminate the novel H1N1 from other swine and human H1 subtype viruses was also developed.

RESULTS: All of the assays had detection limits for the positive control in the range of 1.0 x 10^-4 to 2.0 x 10^-3 of the median tissue culture infective dose. Assay specificities were high, and, for the conventional and real-time assays all negative control samples were negative including 7 human seasonal H1N1 viruses, 1 human H2N2 virus, 2 human seasonal H3N2 viruses, 1 human H5N1 virus, 7 avian influenza viruses (HA subtypes 4, 5, 7, 8, 9, and 10) and 48 nasopharyngeal aspirates (NPAs) from patients with noninfluenza respiratory diseases; for the assay which discriminates the novel H1N1 from other swine and human H1 subtype viruses, all negative controls were also negative, including 20 control NPAs, 2 seasonal human H1N1 viruses, 2 seasonal human H3N2 viruses, and 2 human H5N1 viruses.

CONCLUSIONS: These assays appear useful for the rapid diagnosis of cases with the novel H1N1 virus, thereby allowing better pandemic preparedness.