BACKGROUND: Influenza B viruses belong to two antigenically and genetically distinct lineages which co-circulate in varying proportions in many countries. OBJECTIVE: To develop simple, rapid, accurate and robust methods to detect and differentiate currently circulating B-lineage viruses in respiratory samples and virus isolates. STUDY DESIGN: Haemagglutinin (HA) gene sequences from more than 6300 influenza B strains were analysed to identify signature sequences that could be used to distinguish between B-lineages and sublineages. RESULTS: Pyrosequencing and a real time PCR assays were developed to detect the major B-lineages (B/Victoria/2/87 or B/Yamagata/16/88) and pyrosequencing for a unique mutation was used to further differentiate the B/Yamagata viruses into two currently co-circulating subgroups. More than 300 influenza virus-containing samples, including original specimens, cell and egg grown viruses, were tested with a 100% accuracy. Furthermore, when the same PCR primers were used in an rRT-PCR assay, the two lineages could be differentiated by their distinct ranges of melting temperature with an overall accuracy of 99% for 158 samples tested. CONCLUSIONS: These new pyrosequencing and rRT-PCR methods have the potential to aid the rapid identification of influenza B-lineages for surveillance purposes and to increase the available data for bi-annual selection of viruses for updating influenza vaccines.
BACKGROUND:Influenza B viruses belong to two antigenically and genetically distinct lineages which co-circulate in varying proportions in many countries. OBJECTIVE: To develop simple, rapid, accurate and robust methods to detect and differentiate currently circulating B-lineage viruses in respiratory samples and virus isolates. STUDY DESIGN: Haemagglutinin (HA) gene sequences from more than 6300 influenza B strains were analysed to identify signature sequences that could be used to distinguish between B-lineages and sublineages. RESULTS: Pyrosequencing and a real time PCR assays were developed to detect the major B-lineages (B/Victoria/2/87 or B/Yamagata/16/88) and pyrosequencing for a unique mutation was used to further differentiate the B/Yamagata viruses into two currently co-circulating subgroups. More than 300 influenza virus-containing samples, including original specimens, cell and egg grown viruses, were tested with a 100% accuracy. Furthermore, when the same PCR primers were used in an rRT-PCR assay, the two lineages could be differentiated by their distinct ranges of melting temperature with an overall accuracy of 99% for 158 samples tested. CONCLUSIONS: These new pyrosequencing and rRT-PCR methods have the potential to aid the rapid identification of influenza B-lineages for surveillance purposes and to increase the available data for bi-annual selection of viruses for updating influenza vaccines.
Authors: Dhanasekaran Vijaykrishna; Edward C Holmes; Udayan Joseph; Mathieu Fourment; Yvonne C F Su; Rebecca Halpin; Raphael T C Lee; Yi-Mo Deng; Vithiagaran Gunalan; Xudong Lin; Timothy B Stockwell; Nadia B Fedorova; Bin Zhou; Natalie Spirason; Denise Kühnert; Veronika Bošková; Tanja Stadler; Anna-Maria Costa; Dominic E Dwyer; Q Sue Huang; Lance C Jennings; William Rawlinson; Sheena G Sullivan; Aeron C Hurt; Sebastian Maurer-Stroh; David E Wentworth; Gavin J D Smith; Ian G Barr Journal: Elife Date: 2015-01-16 Impact factor: 8.140
Authors: Svetlana V Shcherbik; Nicholas C Pearce; Marnie L Levine; Alexander I Klimov; Julie M Villanueva; Tatiana L Bousse Journal: PLoS One Date: 2014-03-19 Impact factor: 3.240