Detection of H9 subtype influenza virus by loop-mediated isothermal amplification

Objective:To establish loop-mediated isothermal amplification(LAMP) method for detection of H9 subtype influenza A virus.Methods:The H9 hemagglutinin gene sequence information was obtained from Genbank database.According to alignment analysis of HA gene sequences,a set of specific primers were designed by the LAMP primer design software(PrimerExplorer V4).The gene of H9 hemagglutinin was cloned and served as the positive standard and the reaction system was established and optimized.Then the sensitivity and specificity of the established method were evaluated.Results:The sensitivity of LAMP method was 100 fold higher than that of conventional PCR,with the detection limit of five copies per reaction.There was no crossing-reaction with other subtype influenza virus including H1,H3 and H5.These results suggest that the present LAMP system provided a new avenue for the recognition of H9 subtype virus.Conclusion:The developed LAMP method is a sensitive and reliable assay for quick detection of H9 subtype influenza virus.It is very suitable for the labs at grass root level.