Qualitative detection of avian influenza A (H5N1) viruses: a comparative evaluation of four real-time nucleic acid amplification methods.

Abstract

The aim of this study was to determine the performance of real-time amplification based methods - NASBA, TaqMan, RT-FRET, and RT-PCR LUXtrade mark formats - for the detection of influenza A (H5N1) virus RNA. In an analysis of 54 samples obtained from a range of animal species in Thailand during the period 2003-2006, results showed that the NASBA (H5=98.2%, N1=96.3%), TaqMan (H5=98.2%, N1=96.3%) and FRET (H5=98.2%, N1=96.3%) had significantly higher rates of positive detection than LUX (H5=94.4%, N1=50.0%; P<0.001) for influenza A, H5 and N1 isolates. There were no false-positive results from any methods used in the negative-control group of samples. The limits of analytical detection were at least 10copies/reaction in real-time NASBA and LUX assays, while FRET and TaqMan assay appeared to be less sensitive at > or =100copies/reaction. The assays were relatively specific without cross-reactivity to a number of other influenza strains or viral pathogens. In conclusion, our study demonstrated that real-time NASBA, TaqMan and FRET assays can be used to detect influenza A (H5N1) from a wide range of hosts, and be specific for H5N1 samples obtained during different outbreaks (2003-2006). All assays provided the benefit of rapid influenza H5N1 identification for early diagnosis, in the range of hours, and they are well suited to high throughput analyses.

• Authors: Wasun Chantratita, Chonlaphat Sukasem, Supaporn Kaewpongsri, Chutatip Srichunrusami, Wantanit Pairoj, Arunee Thitithanyanont, Kridsada Chaichoune, Parntep Ratanakron, Thaweesak Songserm, Sudarat Damrongwatanapokin, Olfert Landt
• Journal: Molecular and cellular probes (Mol Cell Probes) 2008 Oct-Dec Vol. 22 Issue 5-6 Pg. 287-93 ISSN: 0890-8508 [Print] England
• PMID: 18672052 (Publication Type: Evaluation Study, Journal Article)

Topics

• Animals
• Birds (microbiology)
• Humans
• Influenza A Virus, H5N1 Subtype (genetics)
• Influenza in Birds (epidemiology, microbiology)
• Influenza, Human (epidemiology, microbiology)
• Nucleic Acid Amplification Techniques (methods)
• Reproducibility of Results
• Sensitivity and Specificity
• Thailand