The purpose of this study was to determine which regions of the VP6 protein
of the murine rotavirus strain EDIM are able to elicit protection against
rotavirus shedding in the adult mouse model following intranasal (i.n.) imm
unization with fragments of VP6 and a subsequent oral EDIM challenge. In th
e initial experiment, the first (fragment AB), middle (BC), or last (CD) pa
rt of VP6 that was genetically fused to maltose-binding protein (MBP) and e
xpressed in Escherichia coli was examined. Mice (BALB/c) immunized with two
9-mug doses of each of the chimeras and 10 mug of the mucosal adjuvant LT(
R192G) were found to be protected against EDIM shedding (80, 92, and nearly
100% reduction, respectively; P less than or equal to 0.01) following chal
lenge. Because CD produced almost complete protection, we prepared four E.
coli-expressed, MBP-fused chimeras containing overlapping fragments of the
CD region (i.e., CD1, CD2, CD3, and CD4) whose lengths ranged from 61 to 67
amino acid residues. Following i.n. immunization, CD1, CD2, and CD4 induce
d significant (P less than or equal to 0.004) protection (88, 84, and 92% r
eduction, respectively). In addition, 11 peptides (18 to 30 residues) of th
e CD region with between 0 and 13 overlapping amino acids were synthesized.
Two 50-mug doses of each peptide with LT(R192G) were administered i.n. to
BALB/c mice. Five peptides were found to elicit significant (P less than or
equal to 0.02) protection. Moreover, a 14-amino-acid region within peptide
6 containing a putative CD4(+) T-cell epitope was found to confer nearly c
omplete protection, suggesting a protective role for CD4(+) T cells. Mice t
hat were protected by fragments BC and CD1 and four of the five protective
synthetic peptides did not develop measurable rotavirus antibodies in serum
or stool, implying that protection induced by these domains was not depend
ent on antibody. Together, these observations suggest that multiple regions
of VP6 can stimulate protection, a region of VP6 as small as 14 amino acid
s containing a CD4(+) T-cell epitope can stimulate nearly complete protecti
on, and protection mediated by a subset of epitopes in the VP6 protein does
not require antibodies in BALB/c mice.