Abstract:
Sequence variation within the 18S SSU rRNA V4 hyper-variable region can affect the accuracy of real-time hybridization
probe-based diagnostics for the detection of Theileria spp. infections. This is relevant for assays that use non-specific
primers, such as the real-time hybridization assay for T. parva (Sibeko et al. 2008). To assess the effect of sequence variation
on this test, the Theileria 18S gene from 62 buffalo and 49 cattle samples was cloned and*1000 clones sequenced. Twentysix
genotypes were detected which included known and novel genotypes for the T. buffeli, T. mutans, T. taurotragi and
T. velifera clades. A novel genotype related to T. sp. (sable) was also detected in 1 bovine sample. Theileria genotypic
diversity was higher in buffalo compared to cattle. Polymorphism within the T. parva hyper-variable region was confirmed
by aberrant real-time melting peaks and supported by sequencing of the S5 ribosomal gene. Analysis of the S5 gene suggests
that this gene can be a marker for species differentiation. T. parva, T. sp. (buffalo) and T. sp. (bougasvlei) remain the only
genotypes amplified by the primer set of the hybridization assay. Therefore, the 18S sequence diversity observed does not
seem to affect the current real-time hybridization assay for T. parva.
