Heartwater, or cowdriosis, is a tick-borne disease of domestic and wild ruminants that is
endemic in the Caribbean and sub-Saharan Africa. The disease is caused by an intracellular
pathogen, Ehrlichia ruminantium and may be fatal within days of the onset of clinical signs
with mortality rates of up to 90% in susceptible hosts. Due to the presence of competent tick
vectors in North America, there is substantial risk of introduction of heartwater with potentially
devastating consequences to the domestic livestock industry. There is currently no reliable
or safe vaccine for use globally. To develop a protective DIVA (differentiate infected
from vaccinated animals) subunit vaccine for heartwater, we targeted the E. ruminantium
immunodominant major antigenic protein1 (MAP1) with the hypothesis that MAP1 is a glycosylated
protein and glycans contained in the antigenic protein are important epitope determinants.
Using a eukaryotic recombinant baculovirus expression system, we expressed and
characterized, for the first time, a glycoform profile of MAP1 of two Caribbean E. ruminantium
isolates, Antigua and Gardel. We have shown that the 37±38 kDa protein corresponded
to a glycosylated form of the MAP1 protein, whereas the 31±32 kDa molecular weight band
represented the non-glycosylated form of the protein frequently reported in scientific literature.
Three groups of sheep (n = 3±6) were vaccinated with increasing doses of a bivalent
(Antigua and Gardel MAP1) rMAP1 vaccine cocktail formulation with montanide ISA25 as
an adjuvant. The glycosylated recombinant subunit vaccine induced E. ruminantium-specific
humoral and Th1 type T cell responses, which are critical for controlling intracellular
pathogens, including E. ruminantium, in infected hosts. These results provide an important
basis for development of a subunit vaccine as a novel strategy to protect susceptible livestock
against heartwater in non-endemic and endemic areas.