Section of Infectious Diseases, Department of Medical Sciences, Uppsala University, Uppsala, Sweden.
PLoS One. 2011;6(9):e24742. doi: 10.1371/journal.pone.0024742. Epub 2011 Sep 12.
Oseltamivir (Tamiflu®) is the most widely used drug against influenza infections and is extensively stockpiled worldwide as part of pandemic preparedness plans. However, resistance is a growing problem and in 2008-2009, seasonal human influenza A/H1N1 virus strains in most parts of the world carried the mutation H274Y in the neuraminidase gene which causes resistance to the drug. The active metabolite of oseltamivir, oseltamivir carboxylate (OC), is poorly degraded in sewage treatment plants and surface water and has been detected in aquatic environments where the natural influenza reservoir, dabbling ducks, can be exposed to the substance. To assess if resistance can develop under these circumstances, we infected mallards with influenza A/H1N1 virus and exposed the birds to 80 ng/L, 1 µg/L and 80 µg/L of OC through their sole water source. By sequencing the neuraminidase gene from fecal samples, we found that H274Y occurred at 1 µg/L of OC and rapidly dominated the viral population at 80 µg/L. IC₅₀ for OC was increased from 2-4 nM in wild-type viruses to 400-700 nM in H274Y mutants as measured by a neuraminidase inhibition assay. This is consistent with the decrease in sensitivity to OC that has been noted among human clinical isolates carrying H274Y. Environmental OC levels have been measured to 58-293 ng/L during seasonal outbreaks and are expected to reach µg/L-levels during pandemics. Thus, resistance could be induced in influenza viruses circulating among wild ducks. As influenza viruses can cross species barriers, oseltamivir resistance could spread to human-adapted strains with pandemic potential disabling oseltamivir, a cornerstone in pandemic preparedness planning. We propose surveillance in wild birds as a measure to understand the resistance situation in nature and to monitor it over time. Strategies to lower environmental levels of OC include improved sewage treatment and, more importantly, a prudent use of antivirals.
奥司他韦（达菲）是最广泛使用的抗流感感染药物，作为大流行准备计划的一部分，在全球范围内广泛储备。然而，耐药性是一个日益严重的问题，2008-2009 年，世界大部分地区的季节性人感染甲型 H1N1 流感病毒株在神经氨酸酶基因中携带 H274Y 突变，导致对该药物的耐药性。奥司他韦的活性代谢物奥司他韦羧酸（OC）在污水处理厂和地表水的降解能力较差，并且已经在水生环境中检测到，那里是可以接触到这种物质的鸭类等自然流感宿主。为了评估在这些情况下是否会产生耐药性，我们用甲型 H1N1 病毒感染野鸭，并通过它们唯一的水源使鸟类暴露于 80ng/L、1μg/L 和 80μg/L 的 OC 下。通过对粪便样本中的神经氨酸酶基因进行测序，我们发现 H274Y 出现在 1μg/L 的 OC 中，并在 80μg/L 的 OC 中迅速占据了病毒群体的主导地位。通过神经氨酸酶抑制测定，OC 对野生型病毒的 IC₅₀ 从 2-4 nM 增加到 H274Y 突变体的 400-700 nM。这与在携带 H274Y 的人类临床分离株中观察到的对 OC 的敏感性降低一致。在季节性爆发期间，OC 的环境水平已测量到 58-293 ng/L，预计在大流行期间将达到μg/L 水平。因此，在野生鸭中循环的流感病毒可能会产生耐药性。由于流感病毒可以跨越物种屏障，奥司他韦耐药性可能会传播到具有大流行潜力的人类适应株，从而使奥司他韦失效，这是大流行准备计划的基石。我们建议对野生鸟类进行监测，以了解自然界中的耐药情况，并随着时间的推移进行监测。降低 OC 环境水平的策略包括改进污水处理，更重要的是，谨慎使用抗病毒药物。
