Emergence of new types of Theileria orientalis in Australian cattle and possible cause of theileriosis outbreaks

Abstract

Theileria parasites cause a benign infection of cattle in parts of Australia where they are endemic, but have, in recent years, been suspected of being responsible for a number of outbreaks of disease in cattle near the coast of New South Wales. The objective of this study was to identify and characterize the species of Theileria in cattle on six farms in New South Wales where disease outbreaks have occurred, and compare with Theileria from three disease-free farms in Queensland that is endemic for Theileria. Special reference was made to sub-typing of T. orientalis by type-specific PCR and sequencing of the small subunit (SSU) rRNA gene, and sequence analysis of the gene encoding a polymorphic merozoite/piroplasm surface protein (MPSP) that may be under immune selection. Nucleotide sequencing of SSU rRNA and MPSP genes revealed the presence of four Theileria genotypes: T. orientalis (buffeli), T. orientalis (ikeda), T. orientalis (chitose) and T. orientalis type 4 (MPSP) or type C (SSU rRNA). The majority of animals showed mixed infections while a few showed single infection. When MPSP nucleotide sequences were translated into amino acids, base transition did not change amino acid composition of the protein product, suggesting possible silent polymorphism. The occurrence of ikeda and type 4 (type C) previously not reported to occur and silent mutation is thought to have enhanced parasite evasion of the host immune response causing the outbreak.

Figure 1

Australian map showing sampling locations in two states of Queensland and New South Wales. There were nine farms in total, three in Queensland and six in New South Wales. All the farms were in the coastal eastern part of Australia as indicated in the map.

Figure 2

Phylogenetic relationships among Theileria types isolated in NSW and QLD based on SSU rRNA sequences. The tree was constructed using the neighbor-joining algorithm with molecular distances estimated by the Kimura-2 parameter model. This tree shows chitose, buffeli and ikeda types in 3 different clades. One corresponding sequence each from T. parva (AF013418) and T. annulata (AY524666) served as outgroups. Bootstrap values are shown as percentages at each node based on 1000 replicates. Branch lengths correlate to the number of substitutions inferred according to the scale shown.

Figure 3

Phylogenetic relationships among Theileria isolated in NSW and QLD based on MPSP sequences. The tree was constructed using the neighbor-joining algorithm with molecular distances estimated by the Kimura-2 parameter model; three clades are noted representing chitose, buffeli and ikeda types. T. parva and T. annulata served as outgroup. Bootstrap values are shown as percentages at each node based on 1000 replicates. Branch lengths correlate to the number of substitutions inferred according to the scale shown.