Abstract
BackgroundThe rapid and accurate identification of the H5 and H7 subtypes of avian influenza (AI) virus is an important step for the control and eradication of highly pathogenic AI outbreaks and for the surveillance of AI viruses that have the potential to undergo changes in pathogenicity in poultry and wild birds. Currently, real-time reverse transcription polymerase chain reaction (RRT-PCR) is routinely used for the rapid detection of the H5 and H7 genes, but misidentification is frequent for emergent isolates and viruses isolated from diverse regions due to the high sequence variation among AI viruses.FindingsIn this study, an RRT-PCR method was tested for the detection of matrix, H5 and H7 genes from diverse subtypes of AI viruses and from field samples obtained through AI surveillance in South Korea over the last four years. Both RRT-PCR and conventional experiment (virus isolation using egg inoculation followed by reverse transcription polymerase chain reaction) agreed on the virus-positive samples. And the comparison of the results with 174 clinical samples showed a high level of agreement without decreasing the specificity and sensitivity.ConclusionsThis assay could be useful tool for the rapid detection of AI using the field samples from domestic poultry and wild birds in South Korea, and continuous regional updates is needed to validate primer sets as the AI virus evolves.
Highlights
The rapid and accurate identification of the H5 and H7 subtypes of avian influenza (AI) virus is an important step for the control and eradication of highly pathogenic AI outbreaks and for the surveillance of AI viruses that have the potential to undergo changes in pathogenicity in poultry and wild birds
Influenza A viruses containing the HAs of subtypes H5 and H7 may become highly pathogenic upon introduction into poultry, and highly pathogenic avian influenza (HPAI) viruses causing devastating economic losses
Real-time reverse transcription polymerase chain reaction (RRT-PCR) assays have been developed and are used globally: these assays are used for the rapid detection of H5 in many Asian countries (H5N1 epidemic) and as a screening method in several countries in Europe and North America [8,9,10,11]
Summary
This assay could be useful tool for the rapid detection of AI using the field samples from domestic poultry and wild birds in South Korea, and continuous regional updates is needed to validate primer sets as the AI virus evolves. Previous published forward and reverse primers and probes for the matrix and H5 genes [20,21] were observed the concern for sensitivity to detect low pathogenic avian influenza (LPAI) viruses including H5 subtype isolated in South Korea. The sensitivity and specificity of the RRT-PCR assay were compared with those of VI in embryonating eggs combined with RT-PCR using 15 reference influenza viruses with different HA types, 71 isolates of Korean AI viruses, and 174 fecal or cloacal samples collected during the HPAI outbreak season from 2010 to 2011 in South Korea. The RRT-PCR for the matrix gene detected 98.8% of 86 influenza A viruses of various subtypes and origins and showed high sensitivity (Table 2 and Additional file 1: Table S1). The sensitivity of this new method assay to detecting the M, H5 and H7 genes of AI viruses were 99.5% (186/187), 98.9% (89/90) and 100% (11/11) and
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