Effective Mucosal Immunity to Anthrax: Neutralizing Antibodies and Th Cell Responses Following Nasal Immunization with Protective Antigen

Abstract

Mucosal, but not parenteral, immunization induces immune responses in both systemic and secretory immune compartments. Thus, despite the reports that Abs to the protective Ag of anthrax (PA) have both anti-toxin and anti-spore activities, a vaccine administered parenterally, such as the aluminum-adsorbed anthrax vaccine, will most likely not induce the needed mucosal immunity to efficiently protect the initial site of infection with inhaled anthrax spores. We therefore took a nasal anthrax vaccine approach to attempt to induce protective immunity both at mucosal surfaces and in the peripheral immune compartment. Mice nasally immunized with recombinant PA (rPA) and cholera toxin (CT) as mucosal adjuvant developed high plasma PA-specific IgG Ab responses. Plasma IgA Abs as well as secretory IgA anti-PA Abs in saliva, nasal washes, and fecal extracts were also induced when a higher dose of rPA was used. The anti-PA IgG subclass responses to nasal rPA plus CT consisted of IgG1 and IgG2b Abs. A more balanced profile of IgG subclasses with IgG1, IgG2a, and IgG2b Abs was seen when rPA was given with a CpG oligodeoxynucleotide as adjuvant, suggesting a role for the adjuvants in the nasal rPA-induced immunity. The PA-specific CD4+ T cells from mice nasally immunized with rPA and CT as adjuvant secreted low levels of CD4+ Th1-type cytokines in vitro, but exhibited elevated IL-4, IL-5, IL-6, and IL-10 responses. The functional significance of the anti-PA Ab responses was established in an in vitro macrophage toxicity assay in which both plasma and mucosal secretions neutralized the lethal effects of Bacillus anthracis toxin.

Bacillus anthracisEdema Toxin Acts as an Adjuvant for Mucosal Immune Responses to Nasally Administered Vaccine Antigens

“…Previously described ELISA was used to assess anti-OVA and anti-PA Ab levels in plasma and external secretions (34,35). Briefly, microtiter plates were coated with OVA (1 mg/ml) or PA (5 g/ml).…”

“…Mice were lightly anesthetized and given 12.5 l of vaccine/nostril. Blood and external secretions (fecal extracts, vaginal washes, and saliva) were collected as previously described (34,35).…”

Protection against anthrax by needle-free mucosal immunization with human anthrax vaccine

“…This suggests a potentially protective role of anti-PA antibodies at mucosal surfaces against anthrax spore infection. Mucosal, but not parenteral, immunization with PA induces immune responses in both systemic and secretory immune compartments [20]. Our study demonstrates that intranasal immunizations with BioThrax™ elicit robust mucosal and systemic antibody responses against PA.…”

“…ELISPOT was performed as previously described with modifications [20,24]. Multiscreen-IP plates (Millipore Corp., Billerica, MA) were pre-wetted with 35% ethanol, washed with PBS, and coated with either PA or BSA (Promega, Madison, WI) at 5 μg/ml, and incubated overnight at 4°C.…”

“…Evidence has shown that mucosal, but not parenteral, immunization induces immune responses in both systemic and secretory immune compartments [20]. Thus, a vaccine administered parenterally, such as the BioThrax™ vaccine, most likely would not induce mucosal immunity against inhalational anthrax.…”