In Vitro Generation of Neuraminidase Inhibitor Resistance in A(H5N1) Influenza Viruses
In Vitro Generation of Neuraminidase Inhibitor Resistance in A(H5N1) Influenza Viruses
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DOI：
10.1128/aac.00334-09
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发表时间：
2009-10-01
影响因子：
4.9
通讯作者：
Barr, Ian G.
中科院分区：
文献类型：
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作者：
Hurt, Aeron C.;Holien, Jessica K.;Barr, Ian G.
为了确定在神经氨酸酶抑制剂选择压力下高致病性A（H5N1）病毒可能出现的突变，在增加奥司他韦或扎那米韦水平的情况下，对两种抗原性不同的毒株进行了连续传代。在奥司他韦压力下，两种A（H5N1）病毒都发生了H274Y神经氨酸酶突变，尽管在一株中该突变与I222M神经氨酸酶突变同时发生。H274Y神经氨酸酶突变显著降低了对奥司他韦的敏感性（与野生型相比，降低了900- 2500倍）。然而，双H274Y/I222M神经氨酸酶突变对耐药性的影响更大，对奥司他韦的敏感性进一步显著降低（与野生型相比降低了8000倍）。当通过反向遗传学将H274Y/I222M双突变引入重组季节性甲型H1N1病毒时，以及在甲型H5N1和甲型H1N1病毒中产生与H274Y结合的I222替代（I222V）时，观察到对奥司他韦敏感性的类似影响。这些病毒仍然对扎那米韦完全敏感，但对帕拉米韦的敏感性有所降低。在存在扎那米韦的情况下传代A（H5N1）病毒后，毒株发生了D198G神经氨酸酶突变，降低了对扎那米韦和奥司他韦的易感性，还发生了E119G神经氨酸酶突变，显示了对扎那米韦的易感性显著降低（与野生型相比降低了1400倍）。在许多耐药菌株中也检测到与受体结合有关的血凝素残基突变。这项研究确定了在奥司他韦或扎那米韦选择压力下甲型H5N1病毒可能出现的突变，以及双神经氨酸酶突变可能导致药物敏感性显著降低。
To identify mutations that can arise in highly pathogenic A(H5N1) viruses under neuraminidase inhibitor selective pressure, two antigenically different strains were serially passaged with increasing levels of either oseltamivir or zanamivir. Under oseltamivir pressure, both A(H5N1) viruses developed a H274Y neuraminidase mutation, although in one strain the mutation occurred in combination with an I222M neuraminidase mutation. The H274Y neuraminidase mutation reduced oseltamivir susceptibility significantly (900- to 2,500-fold compared to the wild type). However the dual H274Y/I222M neuraminidase mutation had an even greater impact on resistance, with oseltamivir susceptibility reduced significantly further (8,000-fold compared to the wild type). A similar affect on oseltamivir susceptibility was observed when the dual H274Y/I222M mutations were introduced, by reverse genetics, into a recombinant seasonal human A(H1N1) virus and also when an alternative I222 substitution (I222V) was generated in combination with H274Y in A(H5N1) and A(H1N1) viruses. These viruses remained fully susceptible to zanamivir but demonstrated reduced susceptibility to peramivir. Following passage of the A(H5N1) viruses in the presence of zanamivir, the strains developed a D198G neuraminidase mutation, which reduced susceptibility to both zanamivir and oseltamivir, and also an E119G neuraminidase mutation, which demonstrated significantly reduced zanamivir susceptibility (1,400-fold compared to the wild type). Mutations in hemagglutinin residues implicated in receptor binding were also detected in many of the resistant strains. This study identified the mutations that can arise in A(H5N1) under either oseltamivir or zanamivir selective pressure and the potential for dual neuraminidase mutations to result in dramatically reduced drug susceptibility.