Leong NKC, Chu DKW, Chu JTS, et al. A Six-Plex Droplet Digital RT-PCR Assay for Seasonal Influenza Virus Typing, Subtyping, and Lineage Determination. Influenza Other Respir Viruses. 2020;10.1111/irv.1
Background: There are two influenza A subtypes (H1 and H3) and two influenza B lineages (Victoria and Yamagata) that currently co-circulate in humans. In this study, we report the development of a six-plex droplet digital RT-PCR (ddRT-PCR) assay that can detect HA and M segments of influenza A (H1, H3, and M) and influenza B (Yamagata HA, Victoria HA, and M) viruses in a single reaction mixture. It can simultaneously detect six different nucleic acid targets in a ddRT-PCR platform.
Methods: The six-plex ddRT-PCR used in this study is an amplitude-based multiplex assay. The analytical performance of the assay was evaluated. Correlation with standard qRT-PCR methodology was assessed using 55 clinical samples.
Results: The assay has a wide dynamic range, and it has good reproducibility within and between runs. The limit of quantification of each target in this assay ranged from 15 copies/reaction for influenza B Victoria M gene to 45 copies/reaction for influenza B Yamagata M gene. In addition, this assay can accurately quantify each of these targets in samples containing viral RNAs from two different viruses that were mixed in a highly skewed ratio. Typing, subtyping, and lineage differentiation data of 55 tested clinical respiratory specimens were found to be identical to those deduced from standard monoplex qRT-PCR assays.
Conclusions: The six-plex ddRT-PCR test was demonstrated to be highly suitable for detecting dual influenza infection cases. This assay is expected to be a useful diagnostic tool for clinical and research use.
Methods: The six-plex ddRT-PCR used in this study is an amplitude-based multiplex assay. The analytical performance of the assay was evaluated. Correlation with standard qRT-PCR methodology was assessed using 55 clinical samples.
Results: The assay has a wide dynamic range, and it has good reproducibility within and between runs. The limit of quantification of each target in this assay ranged from 15 copies/reaction for influenza B Victoria M gene to 45 copies/reaction for influenza B Yamagata M gene. In addition, this assay can accurately quantify each of these targets in samples containing viral RNAs from two different viruses that were mixed in a highly skewed ratio. Typing, subtyping, and lineage differentiation data of 55 tested clinical respiratory specimens were found to be identical to those deduced from standard monoplex qRT-PCR assays.
Conclusions: The six-plex ddRT-PCR test was demonstrated to be highly suitable for detecting dual influenza infection cases. This assay is expected to be a useful diagnostic tool for clinical and research use.
See Also:
Latest articles in those days:
- mRNA vaccines encoding computationally optimized hemagglutinin elicit protective antibodies against future antigenically drifted H1N1 and H3N2 influenza viruses isolated between 2018-2020 11 hours ago
- Clinical Characteristics of 118 Pediatric Patients With Acute Benign Myositis Associated With Influenza A Virus Infection 12 hours ago
- Avian Influenza A Viruses Modulate the Cellular Cytoskeleton during Infection of Mammalian Hosts 12 hours ago
- Seroprevalence of Avian Influenza A(H5N6) Virus Infection, Guangdong Province, China, 2022 3 days ago
- Clinical Presentation, Risk Factors, and Comparison of Laboratory Diagnostics for Seasonal Influenza Virus Among Cambodians From 2007 to 2020 3 days ago
[Go Top] [Close Window]