| 要約: | Influenza virus infection can result in respiratory illnesses and may contribute to high
rate of morbidity and mortality. In addition to the pandemic influenza, seasonal
influenza remains a major health concern worldwide. To date, vaccination is still the
most effective approach for the prevention and control of influenza. Vaccines are
usually delivered by intramuscular (IM) injection using needle, which causes pain and
possibility of blood transmissible infections. Side effects associated with conventional
vaccines, such as local reactions at the injection site, are among the issues necessitating
alternative modes of vaccine delivery. Currently, there is great interest in developing
mucosal vaccine as an alternative over the conventional IM route. It is suggested that
administration of mucosal vaccines will eliminate the need for needles and skilled
personnel, and these would be a more favorable approach to mass vaccination,
especially of children. One of the current mucosal vaccine strategies is by oral
administration of antigen displayed on Lactococcus lactis. In the present study,
influenza A (H1N1) hemagglutinin 1 (HA1) was surface-displayed on non-recombinant
L. lactis. The objective of the study was to evaluate the non-recombinant L. lactis
ability to induce mucosal immune response and to accord protection against influenza
virus infection in mice. The HA1/L/AcmA recombinant protein was constructed by
fusing HA1 with N-acetylmuramidase (AcmA) binding domain using a single-chain
variable fragment (scFv) peptide linker comprising (Gly4Ser)3. The inclusion of this
peptide linker in the recombinant protein was investigated for its ability to improve the
surface display on L. lactis. Flow cytometry and immunoblotting analysis suggested
that the amount of HA1 bound on L. lactis was improved in the presence of this peptide
linker. The recombinant protein completely agglutinated red blood cells (RBCs),
suggesting that insertion of the scFv peptide linker between HA1 and AcmA binding
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domain did not affect the biological function of HA. The non-recombinant L. lactis
displaying HA1/L/AcmA recombinant protein, LL-HA1/L/AcmA, was administered
into mice orally to evaluate the host immune responses. Mice immunized with LLHA1/L/AcmA developed detectable specific sIgA in faecal extract, small intestine wash,
BAL fluid and nasal fluid. The results obtained suggested that oral immunization of
mice with LL-HA1/L/AcmA elicited mucosal immunity in both the gastrointestinal tract
and the respiratory tract. The protective efficacy of LL-HA1/L/AcmA in immunized
mice against a lethal dose challenge with A/TN/1-560/2009-MA2(H1N1) influenza
virus was also assessed. Upon challenge, the PBS-treated group (control) of mice
showed total body weight loss up to 20%, suggesting high susceptibility to influenza
virus infection. In contrast, 7/8 of mice immunized with LL-HA1/L/AcmA and 6/8 of
mice immunized with HA1/L/AcmA recombinant protein survived. The latter group
however, had severe sickness and total body weight loss when compared to the former
group, suggesting that oral immunization with LL-HA1/L/AcmA resulted in less
morbidity and better survival upon challenge with influenza virus. In conclusion, oral
administration of LL-HA1/L/AcmA in mice induced mucosal immunity and provided
protection against lethal challenge with influenza virus. These results highlight the
potential application of L. lactis as a platform for delivery of influenza virus vaccine.
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