Unique Mitochondrial Single Nucleotide Polymorphisms Demonstrate Resolution Potential to Discriminate Theileria parva Vaccine and Buffalo-Derived Strains

Abstract

Distinct pathogenic and epidemiological features underlie different Theileria parva strains resulting in different clinical manifestations of East Coast Fever and Corridor Disease in susceptible cattle. Unclear delineation of these strains limits the control of these diseases in endemic areas. Hence, an accurate characterization of strains can improve the treatment and prevention approaches as well as investigate their origin. Here, we describe a set of single nucleotide polymorphisms (SNPs) based on 13 near-complete mitogenomes of T. parva strains originating from East and Southern Africa, including the live vaccine stock strains. We identified 11 SNPs that are non-preferentially distributed within the coding and non-coding regions, all of which are synonymous except for two within the cytochrome b gene of buffalo-derived strains. Our analysis ascertains haplotype-specific mutations that segregate the different vaccine and the buffalo-derived strains except T. parva-Muguga and Serengeti-transformed strains suggesting a shared lineage between the latter two vaccine strains. Phylogenetic analyses including the mitogenomes of other Theileria species: T. annulata, T. taurotragi, and T. lestoquardi, with the latter two sequenced in this study for the first time, were congruent with nuclear-encoded genes. Importantly, we describe seven T. parva haplotypes characterized by synonymous SNPs and parsimony-informative characters with the other three transforming species mitogenomes. We anticipate that tracking T. parva mitochondrial haplotypes from this study will provide insight into the parasite's epidemiological dynamics and underpin current control efforts.

Keywords: SNPs; Theileria parva; haplotypes; live vaccine; mitogenomes.

Figure A1

Maximum likelihood phylogeny based on cox1 gene sequences (~1.4 kb). The nucleotide substitution model for the tree constructions as determined by the lowest BIC values was GTR + I + G. Bootstrap values are based on 100 replicates. Sequences from this study are in bold.

Figure 1

Linear map of T. parva mitochondrial genome and alignment showing the distribution of variants (SNPs) across the mitogenome sequences obtained by Sanger sequencing. cox1 and cox3: cytochrome oxidase subunits; cob: Cytochrome b; LSU: large subunit; ITR: Inverted terminal repeat region; SNPs: single nucleotide polymorphisms; vertical markings indicate polymorphisms in respective nucleotide sequence relative to the reference sequence AB499089 above.

Figure 2

Percentage pairwise identity of the three protein-coding genes across the 14 mitogenomes analyzed in this study. The 25th and 75th percentiles are represented by the box limits; lines across the boxes indicate the median; whiskers extend to the maximum and minimum (%) identity values.

Figure 3

Maximum likelihood phylogeny based on (A) near-complete whole mitogenome sequences (~5.8 kb) and (B) cob sequences (~1.1 kb). The nucleotide substitution models for the tree constructions as determined by the lowest Bayesian information (BIC) values were TVM + G and GTR + G, respectively. Bootstrap values are based on 100 replicates. Sequences from this study are in bold.

Figure 4

A median-joining (MJ) parsimony network for the 13 T. parva haplotype mitogenome sequences. Node labels TpMtH1-7 represents the unique haplotypes. Lines between nodes indicate mutation points. Larger and fractionated nodes indicate shared haplotypes with multiple strains, each marked with a different color key, as shown.

Figure 5

Phylogenetic grouping of the seven T. parva haplotypes identified in this study. The neighbor-joining tree is constructed based on the Jukes–Cantor Model using 9 T. parva only SNPs out of 662 informative SNPs extracted from 13-mitogenome sequences alignment that included; T. taurotragi, T. annulata, and T. lestoquardi. Numbers behind the nodes indicate bootstrap values based on 1000 replicates. The positions are relative to the AB499089 T. parva Muguga mitogenome sequence. Orthologous positions in the three other transforming Theileria are shown for comparison.