Heterologous expression of glycosylphosphatidyl (GPI)-anchored vaccine candidates of bovine pathogenic protozoa in Tetrahymena thermophila

MONTES, GUADALUPE; ELGUERO, EUGENIA; TOMAZIC, MARIELA L; RODRIGUEZ, ANABEL E.; NUDEL, CLARA; FLORIN-CHRISTENSEN MÓNICA; SCHNITTGER, LEONHARD

Cartagena

Congreso; Vaccine and Research Development 2016; 2016

Tetrahymena thermophila is a free-living non-pathogenic ciliate that has been shown to be suitable for the recombinant expression of glycosylphosphatidylinositol (GPI)-anchored antigens on its surface. Recent examples are the circumsporozoite protein (CSP) of the human malaria pathogen Plasmodium falciparum and the immobilization antigen (i-antigen) of the fish white-spot disease pathogen Ichthyophthirius multifiliis. Our work focused on the use of T. thermophila as expression platform for the GPI-anchored surface proteins MSA-2c and GP60 that constitute vaccine candidates for the apicomplexan protozoa Babesia bovis and Cryptosporidium parvum, respectively. B. bovis is a tick-transmitted hemoprotozooan causing bovine babesiosis of cattle resulting in substantial economic loss in tropical and subtropical regions worldwide. The ubiquitous Cryptosporidium parvum is responsible for high morbidity and mortality of calves raised in dairy farm settings. In addition, infected caves excrete via feces high numbers of the infective oocyst stage posing a risk of human outbreaks due to contamination of water and food. T. thermophila cells were transformed with msa-2c or gp60 gene-carrying suitable vectors for episomal expression. After culturing of recombinant clones, these antigens could be detected in whole cell extracts and enriched membrane fractions by western blot analysis using specific antibodies. Furthermore, the identity of the expressed antigens was verified by HPLC-MS. These results demonstrate that T. thermophila is able to express successfully both these vaccine candidates. In subsequent experiments, MSA2c and GP60 integrative recombinants of T. tetrahymena for stable expression will be generated. (Supported by FonCyt: PICT-2013-1708 and PICT 2012-0695; INTA: PNBIO-1131034 and PNSA-1115053; Fundación Universidad de Morón PID8-2015).