LOW SPECIFICITY OF A NESTED REVERSE TRANSCRIPTION POLYMERASE CHAIN REACTION TO DETECT AVIAN INFLUENZA VIRUS NUCLEOPROTEIN GENE

Abstract

Reverse transcription polymerase chain reaction (RT-PCR) was used routinely to detect the avian influenza virus (AIV) nucleoprotein (NP) gene. The purpose of the present study was to compare the correctness of a nested RT-PCR (nRT-PCR), one conventional RT-PCR with its outer primer (oRT-PCR) and the other conventional RT-PCR with its inner primer (iRT-PCR) to detect AIV NP gene. A total of 365 AI-free fecal swabs (73 pools), 7 tracheal swabs and anllantoic fluid from 25 chicken embryos were used to determine the analytic specificities of those tests. Compared with the iRT-PCR, the nRT-PCR was more sensitive for AIV detection. However, the specificities of nRT-PCR, oRT-PCR and iRT-PCR were 48.6% (35/72), 100% (67/67) and 91.3% (84/92), respectively. The amplifying band was sequenced and confirmed to be the AIV NP gene as the positive control. The specificity of this nRT-PCR is too low to be used for the AIV screening test.