Hippokratia 2012, 16, 4: 384

Gioula G, Melidou A, Exindari M, Chatzidimitriou D, Malisiovas N
National Influenza Centre for northern Greece, Microbiology Department, Medical School, Aristotle University of Thessaloniki


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Keywords:  oseltamivir, influenza, antiviral

Correspoding author: Gioula G, Microbiology Department, Medical School, Aristotle University of Thessaloniki, Greece, e-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

 Dear Editor,

Oseltamivir-resistant infection with the 2009 pandemic influenza A (H1N1) virus has so far been described only rarely and is conferred by the H275Y substitution in the neuraminidase enzyme1. This resistance was unrelated to selective drug pressure, and the H275Y substitution did not appear to reduce transmissibility or severity2.  The purpose of this study was to examine mild, severe and fatal cases of influenza A (H1N1) infections in order to detect strains that possessed the H275Y mutation at the neuraminidase gene during the post pandemic 2010-2011 influenza period.

A total of 50 selective confirmed positive influenza A (H1N1) samples were examined; 16 of those were isolated from fatal cases, 14 from mild and 20 from severe pneumonia cases.  Viral RNA was extracted using the Viral RNA mini kit (Qiagen, Germany) and one step RT-PCR was then performed (Superscript III kit, Invitrogen, UK) with primers amplifying the specific region of the neuraminidase gene. PCR products were purified (Chargeswitch PCR purification, Invitrogen, UK) and then sequenced. Sequences were then analyzed using ClustalW.

Out of the total 50 sequenced neuraminidase genes, 3 were found to possessed  the H275Y mutation. One (6.25%) was isolated from a fatal case and 2 (10%) were isolated from severe pneumonia cases. None of the viruses that caused mild illness possess the mutation.

The fact that no viruses isolated from mild cases possessed the H275Y mutation and that all three resistant strains were isolated from Tamiflu treated cases supports the conclusion that oseltamivir use mainly drives the appearance of the mutation. Compared to the results that we obtained from 84 samples that were analyzed from the pandemic period, when we detected two resistant strains-one was isolated from a mild case and one from a severe pneumonia case-, the present findings lead to the conclusion that the mutation rarely occurs randomly3. However, the overall incidence of the mutation has risen at the post-pandemic period (6%) compared to the pandemic period (2.5%).
According to a similar study in UK, during the winter period 2010/11, 27 confirmed cases of oseltamivir-resistant influenza A (H1N1) 2009 virus infection have been detected. Similar rates of oseltamivir resistance (1%) due to the H275Y mutation were detected in 2010/11 as in 2009/10 period4.

Continuous surveillance for the occurrence of resistant viruses is considered essential, both during and after the upcoming influenza season in the northern hemisphere. Particular attention must be given to detecting transmission of resistant viruses which will mean gathering data on prior antiviral exposure.


1. Updated interim recommendations for the use of antiviral medications in the treatment and prevention of influenza for the 2009-2010 season. Atlanta: Centres for Disease Control and Prevention. Available at:http://www.cdc.gov/h1n1flu/recommendations.htm
2. Oseltamivir-resistant pandemic (H1N1) 2009 influenza virus, October 2009. Wkly Epidemiol Rec. 2009; 84: 453-459.
3. Gioula G, Melidou A, Exindari M, Papoutsi N, Chatzidimitriou D, Dotis J, et al. Oseltamivir-resistant influenza A pandemic (H1N1) 2009 virus in northern Greece. Hippokratia. 2011; 15: 272-274.
4. Lackenby A,  Moran Gilad J, Pebody P, Miah S, Calatayud L, Bolotin S, et al. Continued emergence and changing epidemiology of oseltamivir-resistant influenza A(H1N1)2009 virus, United Kingdom, winter 2010/11. Euro Surveill. 2011; 16: pii=19784.