TY - JOUR
T1 - Whole genome alignment based one-step real-time RT-PCR for universal detection of avian orthoreoviruses of chicken, pheasant and turkey origins
AU - Tang, Yi
AU - Lu, Huaguang
N1 - Funding Information:
This research was funded by the Pennsylvania Department of Agriculture (PDA, ME# 44145029 ), Animal Health and Diagnostic Commission (AHDC)’s research program in 2014–2015, Pennsylvania, USA, The Pennsylvania Poultry Industry Broiler/Egg Check-Off Research Program and The Pennsylvania Soybean Board Research Program , 2015–16, Pennsylvania, USA.
Publisher Copyright:
© 2016 Elsevier B.V.
PY - 2016/4/1
Y1 - 2016/4/1
N2 - Newly emerging avian orthoreovirus (ARV) variants have been continuously detected in Pennsylvania poultry since 2011. In this paper, we report our recent diagnostic assay development of one-step real-time RT-PCR (rRT-PCR) for the rapid and universal detection of all ARVs or reference strains of chicken, pheasant and turkey origins and six σC genotypes of the newly emerging field ARV variants in Pennsylvania (PA) poultry. Primers and probes for the rRT-PCR were designed from the conserved region of the M1 genome segment 5' end based on the whole-genome alignment of various ARV strains, including six field variants or novel strains obtained in PA poultry. The detection limit of the newly developed rRT-PCR for ARV was as low as 10 copies/reaction of viral RNA, and 100.50-100.88 tissue culture infectious dose (TCID50)/100μL of viruses. This new rRT-PCR detected all six σC genotypes from the 66 ARV field variant strains and reference strains tested in this study. There were no cross-reactions with other avian viruses. Reproducibility of the assay was confirmed by intra- and inter-assay tests with variability from 0.12% to 2.19%. Sensitivity and specificity of this new rRT-PCR for ARV were achieved at 100% and 88%, respectively, in comparison with virus isolation as the "gold standard" in testing poultry tissue specimen.
AB - Newly emerging avian orthoreovirus (ARV) variants have been continuously detected in Pennsylvania poultry since 2011. In this paper, we report our recent diagnostic assay development of one-step real-time RT-PCR (rRT-PCR) for the rapid and universal detection of all ARVs or reference strains of chicken, pheasant and turkey origins and six σC genotypes of the newly emerging field ARV variants in Pennsylvania (PA) poultry. Primers and probes for the rRT-PCR were designed from the conserved region of the M1 genome segment 5' end based on the whole-genome alignment of various ARV strains, including six field variants or novel strains obtained in PA poultry. The detection limit of the newly developed rRT-PCR for ARV was as low as 10 copies/reaction of viral RNA, and 100.50-100.88 tissue culture infectious dose (TCID50)/100μL of viruses. This new rRT-PCR detected all six σC genotypes from the 66 ARV field variant strains and reference strains tested in this study. There were no cross-reactions with other avian viruses. Reproducibility of the assay was confirmed by intra- and inter-assay tests with variability from 0.12% to 2.19%. Sensitivity and specificity of this new rRT-PCR for ARV were achieved at 100% and 88%, respectively, in comparison with virus isolation as the "gold standard" in testing poultry tissue specimen.
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U2 - 10.1016/j.meegid.2016.01.018
DO - 10.1016/j.meegid.2016.01.018
M3 - Article
C2 - 26812128
AN - SCOPUS:84955562963
SN - 1567-1348
VL - 39
SP - 120
EP - 126
JO - Infection, Genetics and Evolution
JF - Infection, Genetics and Evolution
ER -
