Abstract:
Low molecular weight (LMW) proteins of E. ruminantium can induce proliferation of
immune peripheral blood mononuclear cells (PBMCs) and the production of interferongamma
(IFN-y ) by CD4+-enriched T-cells. In this study, a reverse vaccinology approach
was applied to identify additional vaccine candidates focusing on genes that encode
LMW proteins smaller than 20 kDa. Five open reading frames (ORFs) were selected from
the E. ruminantium genome and their corresponding recombinant (r) proteins were
produced in a bacterial expression system. Their ability to induce proliferative responses
and IFN-y production was evaluated in vitro using lymphocyte proliferation and
ELISPOT assays. All five recombinant proteins induced proliferation of immune PBMCs
and IFN-y production by these cells. The corresponding five genes were each individually
incorporated into pCMViUBs, a mammalian expression vector and tested as a potential
vaccine in sheep using a DNA prime - protein boost immunisation regimen. A cocktail of
these DNA constructs protected one out of five sheep against a virulent E. ruminantium
(Welgevonden) needle challenge. Three of the five vaccinated sheep showed an increase
in their proliferative responses and production of IFN-y before challenge. This response
decreased after challenge in the sheep that succumbed to the challenge and increased in
the sheep that survived. This finding indicates that sustained IFN -y production is likely to
be involved in conferring protective immunity against heartwater.
