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Background  Pre-pandemic development of an inactivated, split-virion avian influenza vaccine is challenged by the lack of pre-existing immunity and the reduced immunogenicity of some H5 hemagglutinins compared to that of seasonal influenza vaccines. Identification of an acceptable effective adjuvant is needed to improve immunogenicity of a split-virion avian influenza vaccine.    Methods and Findings  Ferrets (N = 118) were vaccinated twice with a split-virion vaccine preparation of A/Vietnam/1203/2004 or saline either 21 days apart (unadjuvanted: 1.9 µg, 7.5 µg, 30 µg, or saline), or 28 days apart (unadjuvanted: 22.5 µg, or alum-adjuvanted: 22.5 or 7.5 µg). Vaccinated animals were challenged intranasally 21 or 28 days later with 10 6  EID 50  of the homologous strain. Immunogenicity was measured by hemagglutination inhibition and neutralization assays. Morbidity was assessed by observed behavior, weight loss, temperature, cytopenias, histopathology, and viral load.  No serum antibodies were detected after vaccination with unadjuvanted vaccine, whereas alum-adjuvanted vaccination induced a robust antibody response. Survival after unadjuvanted dose regimens of 30 µg, 7.5 µg and 1.9 µg (21-day intervals) was 64%, 43%, and 43%, respectively, yet survivors experienced weight loss, fever and thrombocytopenia. Survival after unadjuvanted dose regimen of 22.5 µg (28-day intervals) was 0%, suggesting important differences in intervals in this model. In contrast to unadjuvanted survivors, either dose of alum-adjuvanted vaccine resulted in 93% survival with minimal morbidity and without fever or weight loss. The rarity of brain inflammation in alum-adjuvanted survivors, compared to high levels in unadjuvanted vaccine survivors, suggested that improved protection associated with the alum adjuvant was due to markedly reduced early viral invasion of the ferret brain.    Conclusion  Alum adjuvant significantly improves efficacy of an H5N1 split-virion vaccine in the ferret model as measured by immunogenicity, mortality, morbidity, and brain invasion.

Enhanced Immunogenicity, Mortality Protection, and Reduced Viral Brain Invasion by Alum Adjuvant with an H5N1 Split-Virion Vaccine in the Ferret