Development of a Vaccine Candidate Against Heartwater

The work presented herein reports the improvement achieved on endothelial cell culture conditions and the production of the Rickettsiales Ehrlichia ruminantium (ER) using stirred tanks towards the development of a cost effective process for the widespread application of the inactivated vaccine against heartwater. After the selection of the best endothelial cell line (according to the maximum cell concentration), ER production in stirred tanks was evaluated. Key parameters related with the infection strategy such as serum concentration and multiplicity of infection were then analyzed. The results showed that it was possible to produce ER in stirred tank bioreactors and that the production yields can be increased by a factor of 6 using a serum-free medium during and after the infection process. To expand the process and improve ER production yields, ER growth kinetics was characterized showing that this stirring culture system is capable of efficient bacterial amplification with maximum titers going up to 2.2 log(10)ER/cell. A downstream processing for the purification of ER is also described, taking into account ER recovery yields, the number of steps and endothelial protein contamination. To ensure the successful use of this vaccine, stability tests were also performed with special emphasis on the vaccine formulation and storage temperature; the potency of the final vaccine was validated with test trials in animals (goats). Overall, these results present an attractive process for the production/purification of obligate intracellular bacteria with economical impact in animal and human health, using robust technologies such as stirred tank, microcarriers and membranes.