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Research Project: Development of Detection and Control Strategies for Bovine Babesiosis and Equine Piroplasmosis

Location: Animal Disease Research

Title: The babesia bovis hap2 gene is not required for blood stage replication, but expressed upon in vitro sexual stage induction

Author
item Hussein, H - Washington State University
item Bastos, R - Washington State University
item Schneider, David
item Johnson, W Carl - Carl
item Adham, F - Cairo University
item Davis, W - Washington State University
item Laughery, J - Washington State University
item Herndon, David
item Alzan, H - Washington State University
item Ueti, Massaro
item Suarez, Carlos

Submitted to: PLOS Neglected Tropical Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/14/2017
Publication Date: 10/6/2017
Citation: Hussein, H.E., Bastos, R.G., Schneider, D.A., Johnson, W.C., Adham, F.K., Davis, W.C., Laughery, J.M., Herndon, D.R., Alzan, H.F., Ueti, M.W., Suarez, C.E. 2017. The babesia bovis hap2 gene is not required for blood stage replication, but expressed upon in vitro sexual stage induction. PLOS Neglected Tropical Diseases. 11(10):e0005965. https://doi.org/10.1371/journal.pntd.0005965.
DOI: https://doi.org/10.1371/journal.pntd.0005965

Interpretive Summary: Babesia bovis, is a tick borne apicomplexan parasite responsible for important cattle losses globally. Babesia parasites have a complex life cycle including asexual replication in the mammalian host and sexual reproduction in the tick vector. Novel control strategies aimed at limiting transmission of the parasite are needed, but transmission blocking vaccine candidates remain undefined. In this study we analyze the conservation and role of the hap2 gene in the erythrocyte stage of the life cycle of the parasite and found that expression of the gene is not required for the development of the parasite in erythrocytic stages, using a hap2 mutated parasite line. In addition, we developed an in vitro system for the induction of sexual forms of B. bovis and found expression of the hap2 gene and surface localization of the protein. We concluded that because of the high level of conservation, specific expression in tick stage parasites, and surface exposure, defining HAP2 as a leading candidate for a transmission blocking vaccine against bovine babesiosis.

Technical Abstract: Babesia bovis, is a tick borne apicomplexan parasite responsible for important cattle losses globally. Babesia parasites have a complex life cycle including asexual replication in the mammalian host and sexual reproduction in the tick vector. Novel control strategies aimed at limiting transmission of the parasite are needed, but transmission blocking vaccine candidates remain undefined. Expression of HAP2 has been recognized as critical for the fertilization of parasites in the Babesia-related Plasmodium, and is a leading candidate for a transmission blocking vaccine against malaria. Hereby we identified the B. bovis hap2 gene and demonstrated that it is widely conserved and differentially transcribed during development within the tick midgut, but not by blood stage parasites. The hap2 gene was disrupted by transfecting B. bovis with a plasmid containing the flanking regions of the hap2 gene and the GPF-BSD gene under the control of the ef-1-B promoter. Comparison of in vitro growth between a hap2-KO B. bovis clonal line and its parental wild type strain showed that HAP2 is not required for the development of B. bovis in erythrocytes. However, xanthurenic acid-in vitro induction experiments of sexual stages of parasites recovered after tick transmission resulted in surface expression of HAP2 exclusively in sexual stage induced parasites, as defined both by morphology, and by expression of the B. bovis sexual marker genes 6-Cys A and B. Together, the data strongly suggests that tick midgut stage differential expression of hap2 is associated with the development of B. bovis sexual forms. We concluded that because of the high level of conservation, specific expression in tick stage parasites, and surface exposure, defining HAP2 as a leading candidate for a transmission blocking vaccine against bovine babesiosis.