BACKGROUND: To analyze the genetic and antigenic characteristics of hemagglutinin (HA) gene of human influenza B virus isolated from the mainland of China since 2004-2005. METHODS: The single-way hemagglutinin inhibition (HI) tests were used to test the antigenic characteristics, and the HA1 gene was sequenced based on the antigenic results. RESULTS: The Yamagata-like and Victoria-like viruses co-circulated in 2004-2005. For the Yamagata-like virus, the single-way HI results showed that 3.7% and 4.5% of the viruses had 4-fold greater HI titer difference compared with B/Shanghai/361/02 in 2004 and 2005, respectively. The HA1 sequence data showed that the virus had amino acid mutation, and there was one more glycosylation site at 196th site. For the Victoria-like virus, the single-way HI results showed that 8.5% and 20.6% of the viruses had 4-fold greater HI titer difference compared with B/Hong kong/330/01 in 2004 and 2005, respectively. The HA1 sequence data showed that the virus had replacement of 9 amino acids, and there was one more glycoaylation site at 197th site. CONCLUSION: The results showed that influenza B viruses had changed antigenic and genetic characteristics compared with B/Shanghai/361/02, B/Hong kong/330/01 in 2004-2005.