Development of oseltamivir and zanamivir resistance in influenza A(H1N1)pdm09 virus, Denmark, 2014

Antiviral treatment of immunocompromised patients with prolonged influenza virus infection can lead to multidrug resistance. This study reveals the selection of antiviral resistance mutations in influenza A(H1N1) pdm09 virus in an immunocompromised patient during a 6-month period. The patient was treated with two courses of oseltamivir (5 days and 2 months, respectively), with the first course starting at symptom onset, and subsequently zanamivir (2 months and 10 days, respectively). Respiratory samples were investigated by Sanger and next generation sequencing (NGS) and, for NGS data, low-frequency-variant-detection analysis was performed. Neuraminidase-inhibition tests were conducted for samples isolated in Madin-Darby canine kidney cells. In a sample collected 15 days after the end of the first treatment with oseltamivir (Day 20 post-symptom onset), oseltamivir resistance was detected (mutation H275Y with 60.3% frequency by NGS). Day 149 when the patient had almost completed the second zanamivir treatment, mixes of the following resistance mutations were detected; H275Y(65.1%), I223R(9.2%), and E119G(89.6%), accompanied by additional mutations, showing a more complex viral population in the long-term treated patient. Two samples obtained on Day 151 from bronchoalveolar lavage (BAL) and nasopharyngeal swab, respectively, showed different mutation profiles, with a higher frequency of antiviral resistance mutations in BAL. The results emphasise the importance of timely antiviral resistance testing both for treatment of individual patients as well as for preventive measures to control the development and transmission of antiviral resistant viruses.