Detection of influenza A H1N1 and H3N2 mutations conferring resistance to oseltamivir using rolling circle amplification

作者：Steain Megan C; Dwyer Dominic E; Hurt Aeron C; Kol Chenda; Saksena Nitin K; Cunningham Anthony L; Wang Bin^*

来源：Antiviral Research, 2009, 84(3): 242-248.

DOI：10.1016/j.antiviral.2009.09.010

• Influenza
• Oseltamivir
• Drug resistance
• Rolling circle amplification

In the event of an influenza pandemic, the use of oseltamivir (OTV) will undoubtedly increase and therefore it is more likely that OTV-resistant influenza strains will also arise. OTV-resistance genotyping using sequence-based testing on viruses isolated in cell culture is time consuming and less likely to detect the low-level presence of drug-resistant virus populations. We have developed a novel rolling circle amplification (RCA) method to achieve the sensitive detection of OTV-resistant viruses from clinical specimens. Using artificially created templates, RCA could detect the presence of OTV-resistant mutations (N2: 119V, 292K, N1: 274Y) even if the population carrying the mutations was <1% of the total. By applying RCA to clinical samples, we identified the emergence of the 274Y mutation in one OTV-treated patient, as well as in seven individuals who were treatment-naive (confirming community transmission of 274Y-containing resistant influenza A H1N1). These results were further confirmed by neuraminidase region sequencing. In conclusion, RCA technology can provide rapid (<24h), high-throughput diagnosis of OTV resistance mutations with a high specificity and sensitivity.

• 出版日期2009-12