The identification of Theileria bicornis in captive rhinoceros in Australia

Poaching of both black (Diceros bicornis) and white (Ceratotherium simum) rhinoceros in Africa has increased significantly in recent years. In an effort to ensure the survival of these critically endangered species, breeding programs were established in the 1990s in Australia, where a similar climate and habitat is available. In this study we examined blood samples from two C. simum, including a 16 yr old female (Aluka) who died in captivity, and a 17 yr old asymptomatic male (Umfana). Bloods from seven healthy D. bicornis housed at the zoo were also collected. All samples were tested for the presence of piroplasms via blood smear and PCR. A generic PCR for the 18S rRNA gene of the Piroplasmida revealed the presence of piroplasm infection in both dead and asymptomatic C. simum. Subsequent sequencing of these amplicons revealed the presence of Theileria bicornis. Blood smear indicated that this organism was present at low abundance in both affected and asymptomatic individuals and was not linked to the C. simum mortality. T. bicornis was also detected in the D. bicornis population (n = 7) housed at Taronga Western Plains Zoo using PCR and blood film examination; however only animals imported from Africa (n = 1) tested T. bicornis positive, while captive-born animals bred within Australia (n = 6) tested negative suggesting that transmission within the herd was unlikely. Phylogenetic analysis of the full length T. bicornis 18S rRNA genes classified this organism outside the clade of the transforming and non-transforming Theileria with a new haplotype, H4, identified from D. bicornis. This study revealed the presence of Theileria bicornis in Australian captive populations of both C. simum and D. bicornis and a new haplotype of the parasite was identified. Keywords: Piroplasm, Theileria bicornis, 18S rRNA, Rhinoceros, Translocatio