The increasing number of recent outbreaks of HPAI H5N1 in birds and humans brings out an urgent need to develop potent H5N1 vaccine regimens. Here we present a study on the intranasal vaccination of recombinant baculovirus surface-displayed hemagglutinin (BacHA) or inactivated whole H5N1 viral (IWV) vaccine with a recombinant cholera toxin B subunit (rCTB) as a mucosal adjuvant in a BALB/c mouse model. Two groups of mice were vaccinated with different doses (HA titer of log 2(4) or log 2(8)) of either HA surface-displayed baculovirus or inactivated whole viral vaccine virus adjuvanted with different doses (2 mug or 10 mug) of rCTB. The vaccinations were repeated after 28 days. HA specific serum IgG and mucosal IgA antibodies were quantified by indirect ELISA, and serum neutralizing antibody titer were estimated by hemagglutination inhibition (HI) assay and virus neutralization titer assay. Functional protective efficacy of the vaccine was assessed by host challenge against HPAI H5N1 strains. The results revealed that mice co-administered with log 2(8) HA titer of BacHA vaccine and adjuvanted with 10 mug of rCTB had a significantly enhanced serum IgG and mucosal IgA immune response and serum microneutralization titer compared with mice administered with unadjuvanted log 2(4) or log 2(8) HA titer of BacHA alone. Also vaccination with 10 mug of rCTB and log 2(8) HA titer of BacHA elicited higher HA specific serum and mucosal antibody levels and serum HI titer than vaccination with log 2(8) HA titer of inactivated H5N1 virus adjuvanted with the same dose of rCTB. The host challenge study also showed that 10 mug rCTB combined with log 2(8) HA titer of BacHA provided 100% protection against 10MLD(50) of homologous and heterologous H5N1 strains. The study shows that the combination of rH5 HA expressed on baculovirus surface and rCTB mucosal adjuvant form an effective mucosal vaccine against H5N1 infection. This baculovirus surface-displayed vaccine is more efficacious than inactivated H5N1 influenza vaccine when administered by intranasal route and has no biosafety concerns associated with isolation, purification and production of the latter vaccine.