tetano
Editor, Senior Moderator
Arch Virol. 2019 Feb 21. doi: 10.1007/s00705-019-04180-6. [Epub ahead of print]
[h=1]Multiplex one-step real-time PCR assay for rapid simultaneous detection of velogenic and mesogenic Newcastle disease virus and H5-subtype avian influenza virus.[/h] Zhang Z[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu D[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Hu J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Sun W[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu K[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Li J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Xu H[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], He L[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Jiang D[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Gu M[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Hu S[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Wang X[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu X[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu X[SUP]4,[/SUP][SUP]5,[/SUP][SUP]6[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] H5 avian influenza virus (AIV) and velogenic Newcastle disease virus (v-NDV) are pathogens listed in the OIE Terrestrial Animal Health Code and are considered key pathogens to be eliminated in poultry production. Molecular techniques for rapid detection of H5 AIV and v-NDV are required to investigate their transmission characteristics and to guide prevention. Traditional virus isolation, using embryonated chicken eggs, is time-consuming and cannot be used as a rapid diagnostic technology. In this study, a multiplex real-time RT-PCR (RRT-PCR) detection method for six H5 AIV clades, three v-NDV subtypes, and one mesogenic NDV subtype was successfully established. The detection limit of our multiplex NDV and H5 AIV RRT-PCR was five copies per reaction for each pathogen, with good linearity and efficiency (y = -3.194x + 38.427 for H5 AIV and y = -3.32x + 38.042 for NDV). Multiplex PCR showed good intra- and inter-assay reproducibility, with coefficient of variance (CV) less than 1%. Furthermore, using the RRT-PCR method, H5 AIV and NDV detection rates in clinical samples were higher overall than those obtained using the traditional virus isolation method. Therefore, our method provides a promising technique for surveillance of various H5 AIV clades and multiple velogenic and mesogenic NDV subtypes in live-poultry markets.
PMID: 30790106 DOI: 10.1007/s00705-019-04180-6
[h=1]Multiplex one-step real-time PCR assay for rapid simultaneous detection of velogenic and mesogenic Newcastle disease virus and H5-subtype avian influenza virus.[/h] Zhang Z[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu D[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Hu J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Sun W[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu K[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Li J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Xu H[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu J[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], He L[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Jiang D[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Gu M[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Hu S[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Wang X[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu X[SUP]1,[/SUP][SUP]2,[/SUP][SUP]3[/SUP], Liu X[SUP]4,[/SUP][SUP]5,[/SUP][SUP]6[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] H5 avian influenza virus (AIV) and velogenic Newcastle disease virus (v-NDV) are pathogens listed in the OIE Terrestrial Animal Health Code and are considered key pathogens to be eliminated in poultry production. Molecular techniques for rapid detection of H5 AIV and v-NDV are required to investigate their transmission characteristics and to guide prevention. Traditional virus isolation, using embryonated chicken eggs, is time-consuming and cannot be used as a rapid diagnostic technology. In this study, a multiplex real-time RT-PCR (RRT-PCR) detection method for six H5 AIV clades, three v-NDV subtypes, and one mesogenic NDV subtype was successfully established. The detection limit of our multiplex NDV and H5 AIV RRT-PCR was five copies per reaction for each pathogen, with good linearity and efficiency (y = -3.194x + 38.427 for H5 AIV and y = -3.32x + 38.042 for NDV). Multiplex PCR showed good intra- and inter-assay reproducibility, with coefficient of variance (CV) less than 1%. Furthermore, using the RRT-PCR method, H5 AIV and NDV detection rates in clinical samples were higher overall than those obtained using the traditional virus isolation method. Therefore, our method provides a promising technique for surveillance of various H5 AIV clades and multiple velogenic and mesogenic NDV subtypes in live-poultry markets.
PMID: 30790106 DOI: 10.1007/s00705-019-04180-6