Rethinking Approaches to Improve the Utilization of Nucleic Acid Amplification Tests for Detection and Characterization of Influenza A in Diagnostic and Reference Laboratories

Abstract

Influenza A virus (IFVA) is a significant cause of respiratory infections worldwide and was also responsible for a recent pandemic in 2009. Laboratory identification of IFVA can guide antiviral therapy, assist in cohorting of patients and prevent antibiotic use. Characterization of the virus can track the emergence of novel strains, identify resistance and determine how circulating strains match with vaccine components. The gold standard for detection and characterization of IFVA is nucleic acid amplification technology (e.g., reverse transcriptase PCR [RT-PCR]), which must contend with a constantly evolving viral genome. Although molecular technology has been available for over two decades, there is still an operational gap between assay design and utilization of these tests for the diagnosis and characterization of IFVA. This review will discuss issues surrounding the implementation and use of RT-PCR for the identification and characterization of IFVA, and speculate on why RT-PCR has not been used more widely in clinical laboratories or moved closer to the patient. Newer, less widely used technologies that may change our laboratory practices will be identified and the authors will close with an attempt to identify some future applications of RT-PCR-based technologies for the detection and characterization of IFVA.

Understanding Nucleic Acid Amplification Techniques in the Detection of Influenza viruses in Developing Countries

“…Pyrosequencing is an on-electrophoretic and non-gelbased amplification method that permits sequencing of smaller fragment of sequences (<100 bp) from amplicons generated by RT-PCR [24][25][26][27]. It was developed by Ronaghi et al in 1996 [28], based on enzymatic luminometric inorganic pyrophosphate (PPi) detection assay (ELIDA) established by Nyren in 1987 [29].…”