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Detection and Genogrouping of Noroviruses from Children's Stools By Taqman One-step RT-PCR
Published on: July 22, 2012
A novel real-time RT-PCR assay for influenza C tested in Peruvian children
Leigh M Howard1, Monika Johnson2, Ana I Gil3
1Division of Infectious Diseases, Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN, United States; Vanderbilt Vaccine Research Program, Nashville, TN, United States.
Background:
Influenza C virus (ICV) is associated with acute respiratory illness. Yet ICV remains under recognized, with most previous studies using only culture to identify cases.
Objectives:
To develop a sensitive and specific real-time RT-PCR assay for ICV that allows for rapid and accurate detection in a clinical or research setting.
Study Design:
Multiple ICV sequences obtained from GenBank were analyzed, including 141 hemagglutinin-esterase (HE), 106 matrix (M), and 97 nucleoprotein (NP) sequences. Primers and probes were designed based on conserved regions. Multiple primer-probe sets were tested against multiple ICV strains.
Results:
The ICV M and NP genes offered the most conserved sequence regions. Primers and probes based on newer sequence data offered enhanced detection of ICV, especially for low titer specimens. An NP-targeted assay yielded the best performance and was capable of detecting 10-100 RNA copies per reaction. The NP assay detected multiple clinical isolates of ICV collected in a field epidemiology study conducted in Peru.
Conclusions:
We report a new real-time RT-PCR assay for ICV with high sensitivity and specificity.
