Thompson Catherine I, Barclay Wendy S, Zambon Maria C
School of Animal and Microbial Sciences, University of Reading, Whiteknights PO Box 228, Reading RG6 6AJ.
J Antimicrob Chemother. 2004 May;53(5):759-65. doi: 10.1093/jac/dkh155. Epub 2004 Mar 17.
Influenza A H3N2 viruses isolated recently have characteristic receptor binding properties that may decrease susceptibility to neuraminidase inhibitor drugs. A panel of clinical isolates and recombinant viruses generated by reverse genetics were characterized and tested for susceptibility to zanamivir.
Plaque reduction assays and neuraminidase enzyme inhibition assays were used to assess susceptibility to zanamivir. Receptor binding properties of the viruses were characterized by differential agglutination of red blood cells (RBCs) from different species. Sequence analysis of the haemagglutinin (HA) and neuraminidase (NA) genes was carried out.
Characterization of a panel of H3N2 clinical isolates from 1968 to 2000 showed a gradual decrease in agglutination of chicken and guinea pig RBCs over time, although all isolates could agglutinate turkey RBCs equally. Sequence analysis of the HA and NA genes identified mutations in conserved residues of the HA1 receptor binding site, in particular Leu-226 --> Ile-226/Val-226, and modification of potential glycosylation site motifs. This may be indicative of changes in virus binding to sialic acid (SA) receptors in recent years. Although recent isolates had reduced susceptibility to zanamivir in MDCK cell based plaque reduction assays, no difference was found in an NA enzyme-inhibition assay. Assays with recombinant isogenic viruses showed that the recent HA, but not the NA, conferred reduced susceptibility to zanamivir.
This study demonstrates that recent clinical isolates of influenza A H3N2 virus no longer agglutinate chicken RBCs, but despite significant receptor binding changes as a result of changes in HA, there was little variation in sensitivity of the NA to zanamivir.
近期分离出的甲型H3N2流感病毒具有独特的受体结合特性，这可能会降低其对神经氨酸酶抑制剂药物的敏感性。对一组临床分离株和通过反向遗传学产生的重组病毒进行了特性分析，并检测了它们对扎那米韦的敏感性。
采用蚀斑减少试验和神经氨酸酶抑制试验评估对扎那米韦的敏感性。通过不同物种红细胞（RBC）的差异凝集来表征病毒的受体结合特性。对血凝素（HA）和神经氨酸酶（NA）基因进行序列分析。
对1968年至2000年的一组H3N2临床分离株的特性分析表明，随着时间的推移，鸡和豚鼠红细胞的凝集作用逐渐降低，尽管所有分离株对火鸡红细胞的凝集作用相同。HA和NA基因的序列分析确定了HA1受体结合位点保守残基中的突变，特别是Leu-226→Ile-226/Val-226，以及潜在糖基化位点基序的改变。这可能表明近年来病毒与唾液酸（SA）受体的结合发生了变化。尽管在基于MDCK细胞的蚀斑减少试验中，近期分离株对扎那米韦的敏感性降低，但在NA酶抑制试验中未发现差异。对重组同基因病毒的试验表明，近期的HA而不是NA导致对扎那米韦的敏感性降低。
本研究表明，近期甲型H3N2流感病毒的临床分离株不再凝集鸡红细胞，尽管由于HA的变化导致受体结合发生了显著改变，但NA对扎那米韦的敏感性几乎没有变化。
