Mucosal Immunization with Surface-Displayed Severe Acute Respiratory Syndrome Coronavirus Spike Protein on Lactobacillus casei Induces Neutralizing Antibodies in Mice

다국어 초록 (Multilingual Abstract)

<B>ABSTRACT</B><P>Induction of mucosal immunity may be important for preventing SARS-CoV infections. For safe and effective delivery of viral antigens to the mucosal immune system, we have developed a novel surface antigen display system for lactic acid bacteria using the poly-γ-glutamic acid synthetase A protein (PgsA) of <I>Bacillus subtilis</I> as an anchoring matrix. Recombinant fusion proteins comprised of PgsA and the Spike (S) protein segments SA (residues 2 to 114) and SB (residues 264 to 596) were stably expressed in <I>Lactobacillus casei</I>. Surface localization of the fusion protein was verified by cellular fractionation analyses, immunofluorescence microscopy, and flow cytometry. Oral and nasal inoculations of recombinant <I>L. casei</I> into mice resulted in high levels of serum immunoglobulin G (IgG) and mucosal IgA, as demonstrated by enzyme-linked immunosorbent assays using S protein peptides. More importantly, these antibodies exhibited potent neutralizing activities against severe acute respiratory syndrome (SARS) pseudoviruses. Orally immunized mice mounted a greater neutralizing-antibody response than those immunized intranasally. Three new neutralizing epitopes were identified on the S protein using a peptide neutralization interference assay (residues 291 to 308, 520 to 529, and 564 to 581). These results indicate that mucosal immunization with recombinant <I>L. casei</I> expressing SARS-associated coronavirus S protein on its surface provides an effective means for eliciting protective immune response against the virus.</P>