Dybkaer Karen, Munch Mette, Handberg Kurt J, Jørgensen Poul H
Danish Veterinary Institute, Hangoevej 2, DK-8200 Aarhus N, Denmark.
J Vet Diagn Invest. 2004 Jan;16(1):51-6. doi: 10.1177/104063870401600109.
Three 1-tube Reverse Transcriptase Polymerase Chain Reactions (RT-PCR) directed against the genes encoding the nucleoprotein (NP) and the H5 and H7 hemagglutinin (HA) gene, respectively, were used for detection of avian influenza virus (AIV) in various specimens. A total of 1,040 samples originating from chickens experimentally infected with 2 different low pathogenic avian influenza viruses, from domestic ducks and from wild aquatic birds were examined. The outcome of 1) the universal AIV RT-PCR including a PCR-enzyme-linked immunosorbent assay (ELISA) procedure directed against NP (NP RT-PCR-ELISA) and 2) the subtype specific RT-PCR for H5 and H7 were compared to the results obtained by inoculation of the same specimens into the allantoic cavity of embryonated specific pathogen free (SPF) hen's eggs. Using inoculation in SPF fowl eggs as standard the sensitivity of the NP RT-PCR-ELISA and the RT-PCR for H5 or H7 was 91% and 94%, and the corresponding specificity 98% and 96%. In comparison with inoculation into eggs an additional of 9 samples were positive by NP RT-PCR-ELISA and 13 samples were positive by RT-PCR for one of the HA subtypes. Hence, the 3 RT-PCR procedures described are fast, sensitive and specific for detecting AIV and subtyping H5 and H7 and they are obvious alternatives when testing large numbers of samples.
分别针对编码核蛋白（NP）、H5和H7血凝素（HA）基因的三个单管逆转录聚合酶链反应（RT-PCR）用于检测各种样本中的禽流感病毒（AIV）。共检测了1040份样本，这些样本分别来自实验感染两种不同低致病性禽流感病毒的鸡、家鸭和野生水鸟。将1）通用AIV RT-PCR（包括针对NP的PCR-酶联免疫吸附测定（ELISA）程序，即NP RT-PCR-ELISA）和2）H5和H7亚型特异性RT-PCR的结果与将相同样本接种到无特定病原体（SPF）鸡胚的尿囊腔中所获得的结果进行比较。以接种SPF鸡胚作为标准，NP RT-PCR-ELISA以及H5或H7的RT-PCR的敏感性分别为91%和94%，相应的特异性分别为98%和96%。与接种到鸡蛋中的方法相比，NP RT-PCR-ELISA另外有9个样本呈阳性，RT-PCR针对其中一种HA亚型有13个样本呈阳性。因此，所描述的这三种RT-PCR方法在检测AIV以及对H5和H7进行亚型分型时快速、灵敏且特异，在检测大量样本时是明显的替代方法。
