Transient transfection of Babesia ovis using heterologous promoters

Babesia species, etiological agents of babesiosis, a recognized emerging tick-borne disease, are a significant animal and human health concern with a worldwide socio-economic impact. The development of genetic manipulation techniques, such as transfection technology, is pivotal to improve knowledge regarding the biology of these poorly studied parasites towards better disease control strategies. For Babesia ovis, responsible for ovine babesiosis, a tick-borne disease of small ruminants, these tools are not yet available. The present study was based on the existence of interchangeable cross-species functional promoters between Babesia species. Herein, we describe for the first time B. ovis transient transfection using two heterologous promoters, the ef-1α-B intergenic regions from B. bovis and B. ovata. Their ability to drive expression of a reporter luciferase in B. ovis supports their cross-species functionality. Also, the ef-1α-B promoter region from B. ovata resulted in statistically significantly higher luminescence values in comparison to the control, thus a possibly suitable promoter for stable gene expression. Evaluation of transfection efficiency using qPCR demonstrated that higher luminescence levels were due to promoter strength rather than a higher transfection efficiency. These findings represent a step forward in the development of methods for B. ovis genetic manipulation, an undoubtedly necessary tool to study this parasite basic biology, including its life cycle, the parasite interactions with host cells and virulence factors.