EVALUATION OF THE NucliSENS EasyQ® INFLUENZA A/B ASSAY FOR THE DETECTION OF INFLUENZA A AND B VIRUSES

Objectives: A qualitative real-time NASBA Influenza A/B assay with type specific detection of Influenza A and B RNA in respiratory specimens (nasopharyngeal swabs, washes and aspirates) has been developed and evaluated.

Methods: The analytical limit of detection (95% hit rate) was determined using in vitro RNA transcripts.

To evaluate the biological performances, a total of 430 clinical specimens including 187 nasopharyngeal swabs, 72 nasopharyngeal aspirates and 171 nasal washes has been tested. The reference method was cell culture. Furthermore, QCMD panel has also been evaluated with the kit.

Total nucleic acids were extracted from 200µl of specimen using the NucliSENS easyMAG instrument. Nasopharyngeal washes and aspirates have been ten-fold diluted in UTM-RT before lysis. An Internal Control was added to each sample before extraction and a multiplex real-time NASBA allowed the simultaneously detection of Influenza A, B and Internal Control in a single tube.

Results: The limit of detection of Influenza A/B assay, defined as the 95% hit rate, was found to be 127.0 copies/input [93.8 – 191.0] for Influenza A and 68.6 copies/input [53.6 – 96.7] for Influenza B using in vitro transcripts in extraction.

Out of the 430 clinical specimens, 418 gave valid results. The diagnostic sensitivity was 97.4% against cell culture. The NucliSENS assay was more sensitive than cell culture leading to a diagnostic specificity of 94.7% (14/264 samples found positive in NucliSENS and negative by culture). Real-time NASBA results appeared to be highly correlated with RT-PCR; the diagnostic specificity was therefore at 98% against cell culture combined with RT-PCR results.

The results obtained with the QCMD panel 2007 was 100% in concordance with the expected QCMD results.

Conclusion: With this real-time NASBA assay Influenza A and Influenza B viruses in respiratory specimens can be detected and identified within 3 hours. Results show high sensitivity and specificity and are highly in concordance with culture and RT-PCR.

Molecular biology tests like NucliSENS EasyQ® Influenza A/B are valuable tools alternative to cell culture method for the clinical management of patients with Influenza infections.