Production and purification of chimeric HBc virus-like particles carrying influenza virus LAH domain as vaccine candidates.

BACKGROUND: The lack of a universal influenza vaccine is a global health problem. Interest is now focused on structurally conserved protein domains capable of eliciting protection against a broad range of influenza virus strains. The long alpha helix (LAH) is an attractive vaccine component since it is one of the most conserved influenza hemagglutinin (HA) stalk regions. For an improved immune response, the LAH domain from H3N2 strain has been incorporated into virus-like particles (VLPs) derived from hepatitis B virus core protein (HBc) using recently developed tandem core technology. RESULTS: Fermentation conditions for recombinant HBc-LAH were established in yeast Pichia pastoris and a rapid and efficient purification method for chimeric VLPs was developed to match the requirements for industrial scale-up. Purified VLPs induced strong antibody responses against both group 1 and group 2 HA proteins in mice. CONCLUSION: Our results indicate that the tandem core technology is a useful tool for incorporation of highly hydrophobic LAH domain into HBc VLPs. Chimeric VLPs can be successfully produced in bioreactor using yeast expression system. Immunologic data indicate that HBc VLPs carrying the LAH antigen represent a promising universal influenza vaccine component.

Foot and mouth disease (FMD) virus: quantification of whole virus particles during the vaccine manufacturing process by size exclusion chromatography.

Foot and mouth disease (FMD) is a highly infectious viral disease that affects cattle, sheep, goats and swine causing severe economic losses worldwide. The efficacy of inactivated vaccines is critically dependent on the integrity of foot and mouth disease virus (FMDV) particles. The recommended method to quantify the active ingredient of vaccines is the 140S quantitative sucrose density gradient analysis. This method has been an immensely valuable tool over the past three decades but it is highly operator dependent and difficult to automate. We developed a method to quantify FMDV particles during the vaccine manufacturing process that is based on separation of components by size-exclusion chromatography and measurement of virus by absorption at 254nm. The method is linear in the 5-70µg/mL range, it is applicable to different FMDV strains, and has a good correlation with the 140S test. The proposed method uses standard chromatographic media and it is amenable to automation. The method has potential as a process analytical technology and for control of final product by manufacturers, international vaccine banks and regulatory agencies.

First molecular characterization of canine hepatozoonosis in Argentina: evaluation of asymptomatic Hepatozoon canis infection in dogs from Buenos Aires.

Canine hepatozoonosis is an expanding tick-borne disease in Argentina. Hepatozoonosis was studied during 1 year in six dogs from the same household in Buenos Aires. Blood parasitemia with Hepatozoon gamonts was found in five dogs and all six were positive by PCR for Hepatozoon sp. Although the levels of parasitemia fluctuated during the year, no clinical signs of disease were detected during the follow up period. Amplification and sequencing of a 650 bases fragment of the 18S rRNA gene from all six dogs yielded fragments that were 99% identical to H. canis. The results of the partial 18S rRNA genotyping with the sub-clinical course of infection and lack of severe hematological abnormalities are compatible with clinical and molecular descriptions of Hepatozoon canis infection from other areas of the world. This is the first molecular characterization of Hepatozoon from Argentina.