Okomo-Adhiambo Margaret, Hurt Aeron C, Gubareva Larisa V
Influenza Division, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention, Atlanta, GA, USA.
Methods Mol Biol. 2012;865:95-113. doi: 10.1007/978-1-61779-621-0_6.
Neuraminidase inhibitors (NAIs) represent a newer class of anti-influenza drugs. Widespread natural or acquired resistance to NAIs is a major public health concern as it limits pharmaceutical options available for managing seasonal and pandemic influenza virus infections. Molecular-based methods, such as pyrosequencing, sequencing, and PCR are rapid techniques for detecting known genetic markers of resistance, but they are unable to identify novel mutations that may confer resistance, or subtle differences in the susceptibility of viruses to the NAIs. This chapter describes the chemiluminescent neuraminidase (NA) inhibition (NI) assay, a functional method used for assessing influenza virus susceptibility to NAIs. The assay generates IC(50) values (drug concentration needed to reduce the NA enzymatic activity by 50%) which are determined by curve-fitting analysis. Test viruses showing elevated IC(50) values relative to those of NAI-sensitive reference viruses of the same antigenic type and subtype are further analyzed by pyrosequencing or conventional sequencing to identify known markers of NAI resistance or new changes in the NA. The criteria for NAI resistance are currently not well defined and tend to vary by laboratory and NI assay, therefore harmonization of NI assay conditions and interpretation of results across surveillance laboratories is necessary to improve the NAI susceptibility testing and analysis.
神经氨酸酶抑制剂（NAIs）是一类新型抗流感药物。对NAIs广泛的天然或获得性耐药是一个重大的公共卫生问题，因为它限制了用于控制季节性和大流行性流感病毒感染的药物选择。基于分子的方法，如焦磷酸测序、测序和聚合酶链反应（PCR），是检测已知耐药基因标记的快速技术，但它们无法识别可能赋予耐药性的新突变，或病毒对NAIs敏感性的细微差异。本章描述了化学发光神经氨酸酶（NA）抑制（NI）试验，这是一种用于评估流感病毒对NAIs敏感性的功能方法。该试验产生IC（50）值（将NA酶活性降低50%所需的药物浓度），通过曲线拟合分析确定。相对于相同抗原型和亚型的NAI敏感参考病毒，IC（50）值升高的测试病毒通过焦磷酸测序或常规测序进一步分析，以识别NAI耐药的已知标记或NA中的新变化。目前，NAI耐药的标准尚未明确界定，并且往往因实验室和NI试验而异，因此有必要统一NI试验条件并在各监测实验室之间对结果进行解释，以改进NAI敏感性测试和分析。
