Intratracheal inoculation of AHc vaccine induces protection against aerosolized botulinum neurotoxin A challenge in mice

Abstract

Botulinum neurotoxin (BoNT), produced by Clostridium botulinum, is generally known to be the most poisonous of all biological toxins. In this study, we evaluate the protection conferred by intratracheal (i.t.) inoculation immunization with recombinant Hc subunit (AHc) vaccines against aerosolized BoNT/A intoxication. Three AHc vaccine formulations, i.e., conventional liquid, dry powder produced by spray freeze drying, and AHc dry powder reconstituted in water are prepared, and mice are immunized via i.t. inoculation or subcutaneous (s.c.) injection. Compared with s.c.-AHc-immunized mice, i.t.-AHc-immunized mice exhibit a slightly stronger protection against a challenge with 30,000× LD50 aerosolized BoNT/A. Of note, only i.t.-AHc induces a significantly higher level of toxin-neutralizing mucosal secretory IgA (SIgA) production in the bronchoalveolar lavage of mice. In conclusion, our study demonstrates that the immune protection conferred by the three formulations of AHc is comparable, while i.t. immunization of AHc is superior to s.c. immunization against aerosolized BoNT/A intoxication.

Aerosolized Intratracheal Inoculation of Recombinant Protective Antigen (rPA) Vaccine Provides Protection Against Inhalational Anthrax in B10.D2-Hc0 Mice

“…Formulations containing rPA and CpG were prepared as SFD powder using previously described methods ( 37 ). Briefly, rPA and CpG were dissolved in an aqueous solution of the excipients containing D-mannitol, myo-inositol, L-leucine, and poloxamer 188 at a 1:1 wt/wt ratio.…”

“…Serums and bronchoalveolar lavage fluids (BAL) from four mice per group were collected before each immunization and before the B. anthracis spore challenge. The titers of rPA-specific IgG and SIgA antibodies in serum and mucosal samples were measured by enzyme-linked immunosorbent assay (ELISA) as described previously ( 37 ).They were calculated as the reciprocal of the lowest sample dilution equal to 2.1 times the background optical density (OD) values. Background values were obtained using samples collected from naive mice.…”

“…All measurements were performed in at least triplicate. interferon-γ (IFN-γ) and interleukin-4 (IL-4) levels were measured by enzyme-linked immunospot (ELISPOT) assays as described elsewhere ( 37 ).…”

Complete Protection Against Yersinia pestis in BALB/c Mouse Model Elicited by Immunization With Inhalable Formulations of rF1-V10 Fusion Protein via Aerosolized Intratracheal Inoculation

“…The subunit vaccine powders were prepared by SFD as described in published literature ( 49 , 50 ). Specifically, each subunit vaccine was dissolved in spray drying solution containing D-mannitol, myo-inositol, L-leucine, and poloxamer 188 as excipients and CpG as a mucosal adjuvant ( 51 ).…”

“…The volume median diameter (VMD) of dry powder vaccines was measured using a laser particle size analyzer (RODOS & HELOS, Sympatec, Clausthal-Zellerfeld, Germany) and the mass median aerodynamic diameter (MMAD) of the vaccine aerosol particles was measured with the aerodynamic particle sizer (APS) spectrometer 3321 ( 52 ) (TSI Inc., St. Paul, MN). The moisture content of vaccine dry powders was determined using thermogravimetric analysis (TGA), as described previously ( 50 ). Particle morphology was examined with a Hitachi S-3400N scanning electron microscope.…”