| Summary: | 碩士 === 國立中興大學 === 獸醫學系暨研究所 === 96 === The lactic acid bacteria (LAB) are Gram-positive fermentative microorganisms known primarily for their roles as probiotics. Several lines of evidences have proved that LAB contribute to immunity homeostasis in the intestines and have biological adjuvanticity. Very recently, many studies have focused on developing LAB for the application of mucosal vaccine vehicles. LysM of AcmA’, the autolysin of Lactococcus lactis that could exogenously bind onto the peptidoglycan of LAB, was chosen as an anchor molecule for docking the desire antigenic proteins to LAB. On the basis of literature reviews, four epitopes located at various regions of IBV (Infectious bronchitis virus) spike protein were identified and were successfully expressed as fusion proteins: downstream of each epitopes was tagged with AcmA’ or AcmA’ along with sigmaC (σC), the outer capsid protein of avian reovirus that was chosen for its potential ability of M cells adhesion. Results of whole cell ELISA and western blot analysis have evidenced that the recombinant proteins, σC-AcmA’, AB-AcmA’, EF-AcmA’, GH-σC-AcmA’, and I-σC-AcmA’ were able to bind onto LAB Enterococcus faecium, Lactobacillus salivarius, Lactococcus lactis in trans. The distribution and colonization of LAB were also tested in chicken model; among the three LAB analyzed, E. faecium was the best candidate vector for the high consistence performance between individual animals and was used for further immunization studies. For evaluating the efficacy of LAB-based mucosal vaccination, EF-AcmA’ was adopted as model antigen in combination with or without σC. The results demonstrated mucosal vaccination with EF-AcmA’ bound E. faecium could elicit production of IgA specific for EF epitope in mice. Furthermore, high IgA titer as observed in two out of three mice vaccinated with EF-AcmA’ in the presence of σC, suggesting σC may play a part in immune regulation. However, due to the limited animal size, the properties of immunity enhancement and the M cell targeting of σC should be further characterized.
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